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Sample records for affinity responsive target

  1. Identification of Multiple Cryptococcal Fungicidal Drug Targets by Combined Gene Dosing and Drug Affinity Responsive Target Stability Screening

    Directory of Open Access Journals (Sweden)

    Yoon-Dong Park

    2016-08-01

    Full Text Available Cryptococcus neoformans is a pathogenic fungus that is responsible for up to half a million cases of meningitis globally, especially in immunocompromised individuals. Common fungistatic drugs, such as fluconazole, are less toxic for patients but have low efficacy for initial therapy of the disease. Effective therapy against the disease is provided by the fungicidal drug amphotericin B; however, due to its high toxicity and the difficulty in administering its intravenous formulation, it is imperative to find new therapies targeting the fungus. The antiparasitic drug bithionol has been recently identified as having potent fungicidal activity. In this study, we used a combined gene dosing and drug affinity responsive target stability (GD-DARTS screen as well as protein modeling to identify a common drug binding site of bithionol within multiple NAD-dependent dehydrogenase drug targets. This combination genetic and proteomic method thus provides a powerful method for identifying novel fungicidal drug targets for further development.

  2. Identification of Multiple Cryptococcal Fungicidal Drug Targets by Combined Gene Dosing and Drug Affinity Responsive Target Stability Screening

    Science.gov (United States)

    Park, Yoon-Dong; Sun, Wei; Salas, Antonio; Antia, Avan; Carvajal, Cindy; Wang, Amy; Xu, Xin; Meng, Zhaojin; Zhou, Ming; Tawa, Gregory J.; Dehdashti, Jean; Zheng, Wei; Henderson, Christina M.; Zelazny, Adrian M.

    2016-01-01

    ABSTRACT Cryptococcus neoformans is a pathogenic fungus that is responsible for up to half a million cases of meningitis globally, especially in immunocompromised individuals. Common fungistatic drugs, such as fluconazole, are less toxic for patients but have low efficacy for initial therapy of the disease. Effective therapy against the disease is provided by the fungicidal drug amphotericin B; however, due to its high toxicity and the difficulty in administering its intravenous formulation, it is imperative to find new therapies targeting the fungus. The antiparasitic drug bithionol has been recently identified as having potent fungicidal activity. In this study, we used a combined gene dosing and drug affinity responsive target stability (GD-DARTS) screen as well as protein modeling to identify a common drug binding site of bithionol within multiple NAD-dependent dehydrogenase drug targets. This combination genetic and proteomic method thus provides a powerful method for identifying novel fungicidal drug targets for further development. PMID:27486194

  3. Differential control of Bradyrhizobium japonicum iron stimulon genes through variable affinity of the iron response regulator (Irr) for target gene promoters and selective loss of activator function.

    Science.gov (United States)

    Jaggavarapu, Siddharth; O'Brian, Mark R

    2014-05-01

    Bradyrhizobium japonicum Irr is a conditionally stable transcriptional activator and repressor that accumulates in cells under iron-limited, manganese-replete conditions, but degrades in a haem-dependent manner under high iron conditions, manganese limitation or upon exposure to H2 O2 . Here, we identified Irr-regulated genes that were relatively unresponsive to factors that promote Irr degradation. The promoters of those genes bound Irr with at least 200-fold greater affinity than promoters of the responsive genes, resulting in maintenance of promoter occupancy over a wide cellular Irr concentration range. For Irr-repressible genes, promoter occupancy correlated with transcriptional repression, resulting in differential levels of expression based on Irr affinity for target promoters. However, inactivation of positively controlled genes required neither promoter vacancy nor loss of DNA-binding activity by Irr. Thus, activation and repression functions of Irr may be uncoupled from each other under certain conditions. Abrogation of Irr activation function was haem-dependent, thus haem has two functionally separable roles in modulating Irr activity. The findings imply a greater complexity of control by Irr than can be achieved by conditional stability alone. We suggest that these regulatory mechanisms accommodate the differing needs for Irr regulon genes in response to the prevailing metabolic state of the cell.

  4. Kinetic controlled affinity labeling of target enzyme with thioester chemistry.

    Science.gov (United States)

    Tomohiro, Takenori; Nakabayashi, Masahiro; Sugita, Yuka; Morimoto, Shota

    2016-08-01

    High specificity has been an important feature in affinity labeling for target profiling. Especially, to label targets via rapidly progressing reactions with consumption of ligand (probe), high specificity of reaction with common functional groups of target protein should be achieved without reactions with similar groups of non-target proteins. Herein, we demonstrate the kinetic controlled affinity labeling of acyl CoA synthetase using a fatty acid analogue containing a phenylthioester linkage. High specificity was attained by accelerating the labeling rate in the binding pocket. This approach could be useful for profiling a series of target enzymes and transporters in signal transduction pathways. PMID:27298000

  5. Enhancement of Immune Effector Functions by Modulating IgG’s Intrinsic Affinity for Target Antigen

    Science.gov (United States)

    Mazor, Yariv; Yang, Chunning; Borrok, M. Jack; Ayriss, Joanne; Aherne, Karen; Wu, Herren; Dall’Acqua, William F.

    2016-01-01

    Antibody-mediated immune effector functions play an essential role in the anti-tumor efficacy of many therapeutic mAbs. While much of the effort to improve effector potency has focused on augmenting the interaction between the antibody-Fc and activating Fc-receptors expressed on immune cells, the role of antibody binding interactions with the target antigen remains poorly understood. We show that antibody intrinsic affinity to the target antigen clearly influences the extent and efficiency of Fc-mediated effector mechanisms, and report the pivotal role of antibody binding valence on the ability to regulate effector functions. More particularly, we used an array of affinity modulated variants of three different mAbs, anti-CD4, anti-EGFR and anti-HER2 against a panel of target cell lines expressing disparate levels of the target antigen. We found that at saturating antibody concentrations, IgG variants with moderate intrinsic affinities, similar to those generated by the natural humoral immune response, promoted superior effector functions compared to higher affinity antibodies. We hypothesize that at saturating concentrations, effector function correlates most directly with the amount of Fc bound to the cell surface. Thus, high affinity antibodies exhibiting slow off-rates are more likely to interact bivalently with the target cell, occupying two antigen sites with a single Fc. In contrast, antibodies with faster off-rates are likely to dissociate each binding arm more rapidly, resulting in a higher likelihood of monovalent binding. Monovalent binding may in turn increase target cell opsonization and lead to improved recruitment of effector cells. This unpredicted relationship between target affinity and effector function potency suggests a careful examination of antibody design and engineering for the development of next-generation immunotherapeutics. PMID:27322177

  6. An affinity-directed protein missile system for targeted proteolysis

    Science.gov (United States)

    Fulcher, Luke J.; Macartney, Thomas; Bozatzi, Polyxeni; Hornberger, Annika; Rojas-Fernandez, Alejandro

    2016-01-01

    The von Hippel–Lindau (VHL) protein serves to recruit the hypoxia-inducible factor alpha (HIF1α) protein under normoxia to the CUL2 E3 ubiquitin ligase for its ubiquitylation and degradation through the proteasome. In this report, we modify VHL to engineer an affinity-directed protein missile (AdPROM) system to direct specific endogenous target proteins for proteolysis in mammalian cells. The proteolytic AdPROM construct harbours a cameloid anti-green fluorescence protein (aGFP) nanobody that is fused to VHL for either constitutive or tetracycline-inducible expression. For target proteins, we exploit CRISPR/Cas9 to rapidly generate human kidney HEK293 and U2OS osteosarcoma homozygous knock-in cells harbouring GFP tags at the VPS34 (vacuolar protein sorting 34) and protein associated with SMAD1 (PAWS1, aka FAM83G) loci, respectively. Using these cells, we demonstrate that the expression of the VHL-aGFP AdPROM system results in near-complete degradation of the endogenous GFP-VPS34 and PAWS1-GFP proteins through the proteasome. Additionally, we show that Tet-inducible destruction of GFP-VPS34 results in the degradation of its associated partner, UVRAG, and reduction in levels of cellular phosphatidylinositol 3-phosphate. PMID:27784791

  7. A thermal responsive affinity ligand for precipitation of sialylated proteins

    Directory of Open Access Journals (Sweden)

    Lindsay Arnold

    2016-01-01

    Full Text Available We report here the development of a thermal responsive affinity ligand specific to sialic acid, sialic acid containing oligosaccharides, glycoproteins, and other sialylated glycoconjugates. The ligand is a fusion protein of 40 repeats of pentapeptide of an elastin like polymer (ELP and the 21 kD sialic acid binding domain of a Vibrio cholera neuraminidase (VCNA. For cost-effective synthesis, the fusion protein was targeted to the periplasmic space of an E. coli lpp deletion mutant, resulting in its secretion to the growth medium. A pre-induction heat-shock step at 42 ˚C for 20 minutes was necessary to achieve high level expression of the ligand. Under optimized induction condition (18 ˚C, 0.1 mM IPTG and 48 hours of post-induction cultivation, the ligand was produced to about 100 mg/L. The ligand exhibited a transition temperature of 52 ˚C, which could be depressed to 37 ˚C with the addition of 0.5 M NaCl. Using fetuin as a model sialylated protein, the ligand was applied in an affinity precipitation process to illustrate its potential application in glycoprotein isolation. The ligand captured 100% fetuin from an aqueous solution when the molar ratio of ligand to fetuin was 10 to 1, which was lower than the expected for full titration of sialic acid on the glycoprotein by the lectin. Elution of fetuin from ligand was achieved with PBS buffer containing 2 mM sialic acid. To evaluate how protein and other contaminants influence the recovery of sialylated proteins, CHO medium was spiked into the fetuin solution. The predominant protein species in CHO medium was found to be albumin. Although its removal of over 94% was evident, purified fetuin contained some albumin due to its over-abundance. Additional experiments with albumin contaminant of varying concentrations showed that below 1 mg/L, albumin had no impact on the affinity precipitation, whereas above 10 mg/L, some albumin was co-purified with fetuin. However, even at 50 mg/ml, fetuin

  8. Novel thermo-responsive fucose binding ligands for glycoprotein purification by affinity precipitation.

    Science.gov (United States)

    Arnold, Lindsay; Chen, Rachel

    2014-02-01

    Novel thermo-responsive affinity sugar binders were developed by fusing a bacterial fucose lectin with a thermo-responsive polypeptide. These designer affinity ligand fusions were produced using an Escherichia coli system capable of extracellular secretion of recombinant proteins and were isolated with a high recovery yield (95%) directly from growth medium by Inverse Temperature Cycling (ITC). With horse radish peroxidase (HRP) as a model protein, we demonstrate here that the designer thermo-responsive ligands are capable of interacting with glycans on a glycoprotein, a property that was used to develop a novel affinity precipitation method for glycoprotein purification. The method, requiring only simple process steps, affords full recovery of a target glycoprotein, and is effective at a target glycoprotein concentration as low as 1.4 pM in the presence of large amounts of contaminants. By developing other sugar binders in the similar fashion, the method should be highly useful for glycoprotein purification and detection.

  9. Supramolecular Affinity Chromatography for Methylation-Targeted Proteomics.

    Science.gov (United States)

    Garnett, Graham A E; Starke, Melissa J; Shaurya, Alok; Li, Janessa; Hof, Fraser

    2016-04-01

    Proteome-wide studies of post-translationally methylated species using mass spectrometry are complicated by high sample diversity, competition for ionization among peptides, and mass redundancies. Antibody-based enrichment has powered methylation proteomics until now, but the reliability, pan-specificity, polyclonal nature, and stability of the available pan-specific antibodies are problematic and do not provide a standard, reliable platform for investigators. We have invented an anionic supramolecular host that can form host-guest complexes selectively with methyllysine-containing peptides and used it to create a methylysine-affinity column. The column resolves peptides on the basis of methylation-a feat impossible with a comparable commercial cation-exchange column. A proteolyzed nuclear extract was separated on the methyl-affinity column prior to standard proteomics analysis. This experiment demonstrates that such chemical methyl-affinity columns are capable of enriching and improving the analysis of methyllysine residues from complex protein mixtures. We discuss the importance of this advance in the context of biomolecule-driven enrichment methods. PMID:26973166

  10. GHB receptor targets in the CNS: Focus on high-affinity binding sites

    DEFF Research Database (Denmark)

    Bay, Tina; Eghorn, Laura Friis; Klein, Anders Bue;

    2014-01-01

    γ-Hydroxybutyric acid (GHB) is an endogenous compound in the mammalian brain with both low- and high-affinity receptor targets. GHB is used clinically in the treatment of symptoms of narcolepsy and alcoholism, but also illicitly abused as the recreational drug Fantasy. Major pharmacological effects...... of exogenous GHB are mediated by GABA subtype B (GABAB) receptors that bind GHB with low affinity. The existence of GHB high-affinity binding sites has been known for more than three decades, but the uncovering of their molecular identity has only recently begun. This has been prompted by the generation...... of molecular tools to selectively study high-affinity sites. These include both genetically modified GABAB knock-out mice and engineered selective GHB ligands. Recently, certain GABA subtype A (GABAA) receptor subtypes emerged as high-affinity GHB binding sites and potential physiological mediators of GHB...

  11. GHB receptor targets in the CNS: focus on high-affinity binding sites.

    Science.gov (United States)

    Bay, Tina; Eghorn, Laura F; Klein, Anders B; Wellendorph, Petrine

    2014-01-15

    γ-Hydroxybutyric acid (GHB) is an endogenous compound in the mammalian brain with both low- and high-affinity receptor targets. GHB is used clinically in the treatment of symptoms of narcolepsy and alcoholism, but also illicitly abused as the recreational drug Fantasy. Major pharmacological effects of exogenous GHB are mediated by GABA subtype B (GABAB) receptors that bind GHB with low affinity. The existence of GHB high-affinity binding sites has been known for more than three decades, but the uncovering of their molecular identity has only recently begun. This has been prompted by the generation of molecular tools to selectively study high-affinity sites. These include both genetically modified GABAB knock-out mice and engineered selective GHB ligands. Recently, certain GABA subtype A (GABAA) receptor subtypes emerged as high-affinity GHB binding sites and potential physiological mediators of GHB effects. In this research update, a description of the various reported receptors for GHB is provided, including GABAB receptors, certain GABAA receptor subtypes and other reported GHB receptors. The main focus will thus be on the high-affinity binding targets for GHB and their potential functional roles in the mammalian brain.

  12. Non-affinity factors modulating vascular targeting of nano- and microcarriers.

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    Myerson, Jacob W; Anselmo, Aaron C; Liu, Yaling; Mitragotri, Samir; Eckmann, David M; Muzykantov, Vladimir R

    2016-04-01

    Particles capable of homing and adhering to specific vascular biomarkers have potential as fundamental tools in drug delivery for mediation of a wide variety of pathologies, including inflammation, thrombosis, and pulmonary disorders. The presentation of affinity ligands on the surface of a particle provides a means of targeting the particle to sites of therapeutic interest, but a host of other factors come into play in determining the targeting capacity of the particle. This review presents a summary of several key considerations in nano- and microparticle design that modulate targeted delivery without directly altering epitope-specific affinity. Namely, we describe the effect of factors in definition of the base carrier (including shape, size, and flexibility) on the capacity of carriers to access, adhere to, and integrate in target biological milieus. Furthermore, we present a summary of fundamental dynamics of carrier behavior in circulation, taking into account interactions with cells in circulation and the role of hemodynamics in mediating the direction of carriers to target sites. Finally, we note non-affinity aspects to uptake and intracellular trafficking of carriers in target cells. In total, recent findings presented here may offer an opportunity to capitalize on mitigating factors in the behavior of ligand-targeted carriers in order to optimize targeting. PMID:26596696

  13. High-affinity triplex-forming oligonucleotide target sequences in mammalian genomes.

    Science.gov (United States)

    Wu, Qi; Gaddis, Sara S; MacLeod, Michael C; Walborg, Earl F; Thames, Howard D; DiGiovanni, John; Vasquez, Karen M

    2007-01-01

    Site-specific recognition of duplex DNA by triplex-forming oligonucleotides (TFOs) provides a promising approach to manipulate mammalian genomes. A prerequisite for successful gene targeting using this approach is that the targeted gene must contain specific, high-affinity TFO target sequences (TTS). To date, TTS have been identified and characterized in only approximately 37 human or rodent genes, limiting the application of triplex-directed gene targeting. We searched the complete human and mouse genomes using an algorithm designed to identify high-affinity TTS. The resulting data set contains 1.9 million potential TTS for each species. We found that 97.8% of known human and 95.2% of known mouse genes have at least one potential high-affinity TTS in the promoter and/or transcribed gene regions. Importantly, 86.5% of known human and 83% of the known mouse genes have at least one TTS that is unique to that gene. Thus, it is possible to target the majority of human and mouse genes with specific TFOs. We found substantially more potential TTS in the promoter sequences than in the transcribed gene sequences or intergenic sequences in both genomes. We selected 12 mouse genes and 2 human genes critical for cell signaling, proliferation, and/or carcinogenesis, identified potential TTS in each, and determined TFO binding affinities to these sites in vitro. We identified at least one high-affinity, specific TFO binding site within each of these genes. Using this information, many genes involved in mammalian cell proliferation and carcinogenesis can now be targeted. PMID:17013831

  14. High-affinity triplex targeting of double stranded DNA using chemically modified peptide nucleic acid oligomers

    OpenAIRE

    Hansen, Mads E.; Bentin, Thomas; Nielsen, Peter E.

    2009-01-01

    While sequence-selective dsDNA targeting by triplex forming oligonucleotides has been studied extensively, only very little is known about the properties of PNA–dsDNA triplexes—mainly due to the competing invasion process. Here we show that when appropriately modified using pseudoisocytosine substitution, in combination with (oligo)lysine or 9-aminoacridine conjugation, homopyrimidine PNA oligomers bind complementary dsDNA targets via triplex formation with (sub)nanomolar affinities (at pH 7....

  15. High-affinity triplex targeting of double stranded DNA using chemically modified peptide nucleic acid oligomers

    DEFF Research Database (Denmark)

    Hansen, Mads E; Bentin, Thomas; Nielsen, Peter E

    2009-01-01

    While sequence-selective dsDNA targeting by triplex forming oligonucleotides has been studied extensively, only very little is known about the properties of PNA-dsDNA triplexes-mainly due to the competing invasion process. Here we show that when appropriately modified using pseudoisocytosine...... substitution, in combination with (oligo)lysine or 9-aminoacridine conjugation, homopyrimidine PNA oligomers bind complementary dsDNA targets via triplex formation with (sub)nanomolar affinities (at pH 7.2, 150 mM Na(+)). Binding affinity can be modulated more than 1000-fold by changes in pH, PNA oligomer...... length, PNA net charge and/or by substitution of pseudoisocytosine for cytosine, and conjugation of the DNA intercalator 9-aminoacridine. Furthermore, 9-aminoacridine conjugation also strongly enhanced triplex invasion. Specificity for the fully matched target versus one containing single centrally...

  16. Affinity enhancement of antibodies: how low-affinity antibodies produced early in immune responses are followed by high-affinity antibodies later and in memory B-cell responses.

    Science.gov (United States)

    Eisen, Herman N

    2014-05-01

    The antibodies produced initially in response to most antigens are high molecular weight (MW) immunoglobulins (IgM) with low affinity for the antigen, while the antibodies produced later are lower MW classes (e.g., IgG and IgA) with, on average, orders of magnitude higher affinity for that antigen. These changes, often termed affinity maturation, take place largely in small B-cell clusters (germinal center; GC) in lymphoid tissues in which proliferating antigen-stimulated B cells express the highly mutagenic cytidine deaminase that mediates immunoglobulin class-switching and sequence diversification of the immunoglobulin variable domains of antigen-binding receptors on B cells (BCR). Of the large library of BCR-mutated B cells thus rapidly generated, a small minority with affinity-enhancing mutations are selected to survive and differentiate into long-lived antibody-secreting plasma cells and memory B cells. BCRs are also endocytic receptors; they internalize and cleave BCR-bound antigen, yielding peptide-MHC complexes that are recognized by follicular helper T cells. Imperfect correlation between BCR affinity for antigen and cognate T-cell engagement may account for the increasing affinity heterogeneity that accompanies the increasing average affinity of antibodies. Conservation of mechanisms underlying mutation and selection of high-affinity antibodies over the ≈200 million years of evolution separating bird and mammal lineages points to the crucial role of antibody affinity enhancement in adaptive immunity.

  17. Influence of target concentration and background binding on in vitro selection of affinity reagents.

    Directory of Open Access Journals (Sweden)

    Jinpeng Wang

    Full Text Available Nucleic acid-based aptamers possess many useful features that make them a promising alternative to antibodies and other affinity reagents, including well-established chemical synthesis, reversible folding, thermal stability and low cost. However, the selection process typically used to generate aptamers (SELEX often requires significant resources and can fail to yield aptamers with sufficient affinity and specificity. A number of seminal theoretical models and numerical simulations have been reported in the literature offering insights into experimental factors that govern the effectiveness of the selection process. Though useful, these previous models have not considered the full spectrum of experimental factors or the potential impact of tuning these parameters at each round over the course of a multi-round selection process. We have developed an improved mathematical model to address this important question, and report that both target concentration and the degree of non-specific background binding are critical determinants of SELEX efficiency. Although smaller target concentrations should theoretically offer superior selection outcome, we show that the level of background binding dramatically affect the target concentration that will yield maximum enrichment at each round of selection. Thus, our model enables experimentalists to determine appropriate target concentrations as a means for protocol optimization. Finally, we perform a comparative analysis of two different selection methods over multiple rounds of selection, and show that methods with inherently lower background binding offer dramatic advantages in selection efficiency.

  18. Affinity-based release of polymer-binding peptides from hydrogels with the target segments of peptides.

    Science.gov (United States)

    Serizawa, Takeshi; Fukuta, Hiroki; Date, Takaaki; Sawada, Toshiki

    2016-02-01

    Peptides with affinities for the target segments of polymer hydrogels were identified by biological screening using phage-displayed peptide libraries, and these peptides exhibited an affinity-based release capability from hydrogels. The results from cell culture assays demonstrated the sustained anticancer effects of the drug-conjugated peptides that were released from the hydrogels.

  19. Targeting Protein-Protein Interactions with Trimeric Ligands: High Affinity Inhibitors of the MAGUK Protein Family

    DEFF Research Database (Denmark)

    Nissen, Klaus B; Kedström, Linda Maria Haugaard; Wilbek, Theis S;

    2015-01-01

    related MAGUK proteins contain three consecutive PDZ domains, hence we envisioned that targeting all three PDZ domains simultaneously would lead to more potent and potentially more specific interactions with the MAGUK proteins. Here we describe the design, synthesis and characterization of a series of...... trimeric ligands targeting all three PDZ domains of PSD-95 and the related MAGUK proteins, PSD-93, SAP-97 and SAP-102. Using our dimeric ligands targeting the PDZ1-2 tandem as starting point, we designed novel trimeric ligands by introducing a PDZ3-binding peptide moiety via a cysteine-derivatized NPEG...... linker. The trimeric ligands generally displayed increased affinities compared to the dimeric ligands in fluorescence polarization binding experiments and optimized trimeric ligands showed low nanomolar inhibition towards the four MAGUK proteins, thus being the most potent inhibitors described. Kinetic...

  20. Humanization of high-affinity antibodies targeting glypican-3 in hepatocellular carcinoma

    Science.gov (United States)

    Zhang, Yi-Fan; Ho, Mitchell

    2016-01-01

    Glypican-3 (GPC3) is a cell-surface heparan sulfate proteoglycan highly expressed in hepatocellular carcinoma (HCC). We have generated a group of high-affinity mouse monoclonal antibodies targeting GPC3. Here, we report the humanization and testing of these antibodies for clinical development. We compared the affinity and cytotoxicity of recombinant immunotoxins containing mouse single-chain variable regions fused with a Pseudomonas toxin. To humanize the mouse Fvs, we grafted the combined KABAT/IMGT complementarity determining regions (CDR) into a human IgG germline framework. Interestingly, we found that the proline at position 41, a non-CDR residue in heavy chain variable regions (VH), is important for humanization of mouse antibodies. We also showed that two humanized anti-GPC3 antibodies (hYP7 and hYP9.1b) in the IgG format induced antibody-dependent cell-mediated cytotoxicity and complement-dependent-cytotoxicity in GPC3-positive cancer cells. The hYP7 antibody was tested and showed inhibition of HCC xenograft tumor growth in nude mice. This study successfully humanizes and validates high affinity anti-GPC3 antibodies and sets a foundation for future development of these antibodies in various clinical formats in the treatment of liver cancer. PMID:27667400

  1. Enhanced membrane pore formation through high-affinity targeted antimicrobial peptides.

    Directory of Open Access Journals (Sweden)

    Christopher J Arnusch

    Full Text Available Many cationic antimicrobial peptides (AMPs target the unique lipid composition of the prokaryotic cell membrane. However, the micromolar activities common for these peptides are considered weak in comparison to nisin, which follows a targeted, pore-forming mode of action. Here we show that AMPs can be modified with a high-affinity targeting module, which enables membrane permeabilization at low concentration. Magainin 2 and a truncated peptide analog were conjugated to vancomycin using click chemistry, and could be directed towards specific membrane embedded receptors both in model membrane systems and whole cells. Compared with untargeted vesicles, a gain in permeabilization efficacy of two orders of magnitude was reached with large unilamellar vesicles that included lipid II, the target of vancomycin. The truncated vancomycin-peptide conjugate showed an increased activity against vancomycin resistant Enterococci, whereas the full-length conjugate was more active against a targeted eukaryotic cell model: lipid II containing erythrocytes. This study highlights that AMPs can be made more selective and more potent against biological membranes that contain structures that can be targeted.

  2. Characterization of the ER-Targeted Low Affinity Ca(2+) Probe D4ER.

    Science.gov (United States)

    Greotti, Elisa; Wong, Andrea; Pozzan, Tullio; Pendin, Diana; Pizzo, Paola

    2016-01-01

    Calcium ion (Ca(2+)) is a ubiquitous intracellular messenger and changes in its concentration impact on nearly every aspect of cell life. Endoplasmic reticulum (ER) represents the major intracellular Ca(2+) store and the free Ca(2+) concentration ([Ca(2+)]) within its lumen ([Ca(2+)]ER) can reach levels higher than 1 mM. Several genetically-encoded ER-targeted Ca(2+) sensors have been developed over the last years. However, most of them are non-ratiometric and, thus, their signal is difficult to calibrate in live cells and is affected by shifts in the focal plane and artifactual movements of the sample. On the other hand, existing ratiometric Ca(2+) probes are plagued by different drawbacks, such as a double dissociation constant (Kd) for Ca(2+), low dynamic range, and an affinity for the cation that is too high for the levels of [Ca(2+)] in the ER lumen. Here, we report the characterization of a recently generated ER-targeted, Förster resonance energy transfer (FRET)-based, Cameleon probe, named D4ER, characterized by suitable Ca(2+) affinity and dynamic range for monitoring [Ca(2+)] variations within the ER. As an example, resting [Ca(2+)]ER have been evaluated in a known paradigm of altered ER Ca(2+) homeostasis, i.e., in cells expressing a mutated form of the familial Alzheimer's Disease-linked protein Presenilin 2 (PS2). The lower Ca(2+) affinity of the D4ER probe, compared to that of the previously generated D1ER, allowed the detection of a conspicuous, more clear-cut, reduction in ER Ca(2+) content in cells expressing mutated PS2, compared to controls. PMID:27598166

  3. Characterization of the ER-Targeted Low Affinity Ca2+ Probe D4ER

    Directory of Open Access Journals (Sweden)

    Elisa Greotti

    2016-09-01

    Full Text Available Calcium ion (Ca2+ is a ubiquitous intracellular messenger and changes in its concentration impact on nearly every aspect of cell life. Endoplasmic reticulum (ER represents the major intracellular Ca2+ store and the free Ca2+ concentration ([Ca2+] within its lumen ([Ca2+]ER can reach levels higher than 1 mM. Several genetically-encoded ER-targeted Ca2+ sensors have been developed over the last years. However, most of them are non-ratiometric and, thus, their signal is difficult to calibrate in live cells and is affected by shifts in the focal plane and artifactual movements of the sample. On the other hand, existing ratiometric Ca2+ probes are plagued by different drawbacks, such as a double dissociation constant (Kd for Ca2+, low dynamic range, and an affinity for the cation that is too high for the levels of [Ca2+] in the ER lumen. Here, we report the characterization of a recently generated ER-targeted, Förster resonance energy transfer (FRET-based, Cameleon probe, named D4ER, characterized by suitable Ca2+ affinity and dynamic range for monitoring [Ca2+] variations within the ER. As an example, resting [Ca2+]ER have been evaluated in a known paradigm of altered ER Ca2+ homeostasis, i.e., in cells expressing a mutated form of the familial Alzheimer’s Disease-linked protein Presenilin 2 (PS2. The lower Ca2+ affinity of the D4ER probe, compared to that of the previously generated D1ER, allowed the detection of a conspicuous, more clear-cut, reduction in ER Ca2+ content in cells expressing mutated PS2, compared to controls.

  4. Serum Stability and Affinity Optimization of an M2 Macrophage-Targeting Peptide (M2pep).

    Science.gov (United States)

    Ngambenjawong, Chayanon; Gustafson, Heather H; Pineda, Julio M; Kacherovsky, Nataly A; Cieslewicz, Maryelise; Pun, Suzie H

    2016-01-01

    Tumor associated macrophages (TAMs) are a major stromal component of the tumor microenvironment in several cancers. TAMs are a potential target for adjuvant cancer therapies due to their established roles in promoting proliferation of cancer cells, angiogenesis, and metastasis. We previously discovered an M2 macrophage-targeting peptide (M2pep) which was successfully used to target and deliver a pro-apoptotic KLA peptide to M2-like TAMs in a CT-26 colon carcinoma model. However, the effectiveness of in vivo TAM-targeting using M2pep is limited by its poor serum stability and low binding affinity. In this study, we synthesized M2pep derivatives with the goals of increasing serum stability and binding affinity. Serum stability evaluation of M2pepBiotin confirmed its rapid degradation attributed to exolytic cleavage from the N-terminus and endolytic cleavages at the W10/W11 and S16/K17 sites. N-terminal acetylation of M2pepBiotin protected the peptide against the exolytic degradation while W10w and K(17,18,19)k substitutions were able to effectively protect endolytic degradation at their respective cleavage sites. However, no tested amino acid changes at the W10 position resulted in both protease resistance at that site and retention of binding activity. Therefore, cyclization of M2pep was investigated. Cyclized M2pep better resisted serum degradation without compromising binding activity to M2 macrophages. During the serum stability optimization process, we also discovered that K9R and W10Y substitutions significantly enhanced binding affinity of M2pep. In an in vitro binding study of different M2pep analogs pre-incubated in mouse serum, cyclic M2pep with K9R and W10Y modifications (cyclic M2pep(RY)) retained the highest binding activity to M2 macrophages over time due to its improved serum stability. Finally, we evaluated the in vivo accumulation of sulfo-Cy5-labeled M2pep and cyclic M2pep(RY) in both the CT-26 and 4T1 breast carcinoma models. Cyclic M2pep

  5. Development of immobilized membrane-based affinity columns for use in the online characterization of membrane bound proteins and for targeted affinity isolations

    Energy Technology Data Exchange (ETDEWEB)

    Moaddel, Ruin [Gerontology Research Center, National Institute on Aging, National Institutes of Health, 5600 Nathan Shock Drive, Baltimore, MD 21224-6825 (United States); Wainer, Irving W. [Gerontology Research Center, National Institute on Aging, National Institutes of Health, 5600 Nathan Shock Drive, Baltimore, MD 21224-6825 (United States)]. E-mail: Wainerir@grc.nia.nih.gov

    2006-03-30

    Membranes obtained from cell lines that express or do not express a target membrane bound protein have been immobilized on a silica-based liquid chromatographic support or on the surface of an activated glass capillary. The resulting chromatographic columns have been placed in liquid chromatographic systems and used to characterize the target proteins and to identify small molecules that bind to the target. Membranes containing ligand gated ion channels, G-protein coupled receptors and drug transporters have been prepared and characterized. If a marker ligand has been identified for the target protein, frontal or zonal displacement chromatographic techniques can be used to determine binding affinities (K {sub d} values) and non-linear chromatography can be used to assess the association (k {sub on}) and dissociation (k {sub off}) rate constants and the thermodynamics of the binding process. Membrane-based affinity columns have been created using membranes from a cell line that does not express the target protein (control) and the same cell line that expresses the target protein (experimental) after genomic transfection. The resulting columns can be placed in a parallel chromatography system and the differential retention between the control and experimental columns can be used to identify small molecules and protein that bind to the target protein. These applications will be illustrated using columns created using cellular membranes containing nicotinic acetylcholine receptors and the drug transporter P-glycoprotein.

  6. Affinity ultrafiltration of DNA topoisomerases-targeted compounds determined with HPLC/ESI-MS for drug candidate screening

    Institute of Scientific and Technical Information of China (English)

    张虹; 潘远江

    2004-01-01

    A method of screening assay is demonstrated. The approach is based on the affinity ofantitumor candidates for topoisomerases. In this method, antitumor candidates are fished out using topoisomerases as targets. Traditional analysis of complex compounds typically encounters signal suppression due to the relatively low concentrations, but enzyme-affinity screening for the active compounds can effectively concentrate the desired analysts into a small volume of high concentration. Active compounds are separated from non-affinity compounds by ultrafiltration. The molecules-enzymes complexes that are retained on the filter are subsequently separated by acidification to obtain the topoisomerases-affinity compounds for analysis on High Performance Liquid Chromatography coupled with electrospray ionization mass spectrometric detection (ESI-MS). This enzyme-affinity based screening assay provides a highly specific and efficient method that can directly screen, identify, and acquire drug candidates thus improving the accuracy and speed of high-throughput screening activities.

  7. Affinity ultrafiltration of DNA topoisomerases-targeted compounds determined with HPLC/ESI-MS for drug candidate screening

    Institute of Scientific and Technical Information of China (English)

    张虹; 潘远江

    2004-01-01

    A method of screening assay is demonstrated. The approach is based on the affinity of antitumor candidates for topoisomerases. In this method, antitumor candidates are fished out using topoisomerases as targets. Traditional analysis of complex compounds typically encounters signal suppression due to the relatively low concentrations, but enzyme-affinity screening for the active compounds can effectively concentrate the desired analysts into a small volume of high concen-tration. Active compounds are separated from non-affinity compounds by ultrafiltration. The molecules-enzymes complexes that are retained on the filter are subsequently separated by acidification to obtain the topoisomerases-affinity compounds for analysis on High Performance Liquid Chromatography coupled with electrospray ionization mass spectrometric detec-tion (ESI-MS). This enzyme-affinity based screening assay provides a highly specific and efficient method that can directly screen, identify, and acquire drug candidates thus improving the accuracy and speed of high-throughput screening activities.

  8. The Interplay of Antigen Affinity, Internalization, and Pharmacokinetics on CD44-Positive Tumor Targeting of Monoclonal Antibodies.

    Science.gov (United States)

    Glatt, Dylan M; Beckford Vera, Denis R; Parrott, Matthew C; Luft, J Christopher; Benhabbour, S Rahima; Mumper, Russell J

    2016-06-01

    Monoclonal antibodies (mAbs) offer promise as effective tumor targeting and drug delivery agents for cancer therapy. However, comparative biological and clinical characteristics of mAbs targeting the same tumor-associated antigen (TAA) often differ widely. This study examined the characteristics of mAbs that impact tumor targeting using a panel of mAb clones specific to the cancer-associated cell-surface receptor and cancer stem cell marker CD44. CD44 mAbs were screened for cell-surface binding, antigen affinity, internalization, and CD44-mediated tumor uptake by CD44-positive A549 cells. It was hypothesized that high-affinity, rapidly internalizing CD44 mAbs would result in high tumor uptake and prolonged tumor retention. Although high-affinity clones rapidly bound and were internalized by A549 cells in vitro, an intermediate-affinity clone demonstrated significantly greater tumor uptake and retention than high-affinity clones in vivo. Systemic exposure, rather than high antigen affinity or rapid internalization, best associated with tumor targeting of CD44 mAbs in A549 tumor-bearing mice. PMID:27079967

  9. Affinity capture using peptide-functionalized magnetic nanoparticles to target Staphylococcus aureus

    Science.gov (United States)

    Kuo, Fang-Yin; Lin, Wei-Lien; Chen, Yu-Chie

    2016-04-01

    Staphylococcus aureus, a commonly found pathogen, can cause food poisoning and infections. Thus, it is necessary to develop analytical methods for the rapid screening of S. aureus in suspicious samples. Magnetic nanoparticles (MNPs) are widely used as affinity probes to selectively enrich target species from complex samples because of their high specific surface area and magnetic properties. The MNP surface should be functionalized to have the capability to target specific species. We herein propose a straightforward method to functionalize aluminum oxide-coated iron oxide (Fe3O4@Al2O3) MNPs with the peptide HHHHHHDEEGLFVD (D). The peptide D was comprised of three domains: polyhistidine (H6) used as the linker, DEE added as the spacer, and GLFVD used for targeting S. aureus. D was immobilized on the surface of Fe3O4@Al2O3 MNPs through H6-Al chelation. Our results showed that the D-functionalized Fe3O4@Al2O3 MNPs (D-Fe3O4 MNPs) possess the capability to target S. aureus. The selective trapping experiments were conducted under microwave-heating for only 60 s, and sufficient bacterial cells were trapped by the MNPs to be identified by matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS). We demonstrated that the D-Fe3O4@Al2O3 MNPs combined with MALDI-MS can be used to rapidly characterize trace amounts of S. aureus in complex juice and egg samples.Staphylococcus aureus, a commonly found pathogen, can cause food poisoning and infections. Thus, it is necessary to develop analytical methods for the rapid screening of S. aureus in suspicious samples. Magnetic nanoparticles (MNPs) are widely used as affinity probes to selectively enrich target species from complex samples because of their high specific surface area and magnetic properties. The MNP surface should be functionalized to have the capability to target specific species. We herein propose a straightforward method to functionalize aluminum oxide-coated iron oxide (Fe3O4@Al2O3) MNPs with the

  10. Antibody response and antibody affinity maturation in cats with experimental proliferative immune complex glomerulonephritis.

    Science.gov (United States)

    Bishop, S A; Bailey, M; Lucke, V M; Stokes, C R

    1992-07-01

    An experimental model of proliferative glomerulonephritis (GN) in the cat, which closely resembles human proliferative forms of GN, has been used to study the role of antibody and antibody affinity in the development of immune complex-mediated renal disease. The serum IgG and IgM antibody response to antigen, average antibody affinity (avidity) and affinity heterogeneity of the IgG and IgM populations was assessed at varying times after commencement of chronic immunization with the antigen, human serum albumin (HSA), by enzyme immunoassay. Cats could be classified according to whether they were "low", "intermediate" or "high" IgG responders, by quantification of serum IgG values. Cats with the lowest serum IgG values failed to develop glomerulonephritis. However, there was no relationship between actual IgG values and the severity of the induced disease. In contrast to IgG, there was no division of cats into low or high IgM anti-HSA responders. Again, cats with the lowest IgM values failed to develop GN, but, more interestingly, a late, marked increase in serum IgM anti-HSA occurred only in cats that developed clinical signs of GN (anterior uveitis and nephrotic syndrome). Maturation of average, functional IgG affinity (avidity) for HSA following chronic immunization was clearly demonstrated for all cats. At the end of the experiment, all cats had IgG of high affinity for HSA and the average affinity heterogeneity of the IgG populations was less than in measurements taken earlier. Values of IgG affinity at the end of the experiment were very similar both in cats which developed GN and in those which remained clinically, biochemically and pathologically normal. In contrast to IgG antibody, some cats developed IgM of increased affinity, whilst others produced antibody of reduced affinity, following chronic immunization. There was no correlation between the development of disease and the production of either low or high affinity IgM antibody. Data indicated that an

  11. Viral Escape Mutant Epitope Maintains TCR Affinity for Antigen yet Curtails CD8 T Cell Responses.

    Directory of Open Access Journals (Sweden)

    Shayla K Shorter

    Full Text Available T cells have the remarkable ability to recognize antigen with great specificity and in turn mount an appropriate and robust immune response. Critical to this process is the initial T cell antigen recognition and subsequent signal transduction events. This antigen recognition can be modulated at the site of TCR interaction with peptide:major histocompatibility (pMHC or peptide interaction with the MHC molecule. Both events could have a range of effects on T cell fate. Though responses to antigens that bind sub-optimally to TCR, known as altered peptide ligands (APL, have been studied extensively, the impact of disrupting antigen binding to MHC has been highlighted to a lesser extent and is usually considered to result in complete loss of epitope recognition. Here we present a model of viral evasion from CD8 T cell immuno-surveillance by a lymphocytic choriomeningitis virus (LCMV escape mutant with an epitope for which TCR affinity for pMHC remains high but where the antigenic peptide binds sub optimally to MHC. Despite high TCR affinity for variant epitope, levels of interferon regulatory factor-4 (IRF4 are not sustained in response to the variant indicating differences in perceived TCR signal strength. The CD8+ T cell response to the variant epitope is characterized by early proliferation and up-regulation of activation markers. Interestingly, this response is not maintained and is characterized by a lack in IL-2 and IFNγ production, increased apoptosis and an abrogated glycolytic response. We show that disrupting the stability of peptide in MHC can effectively disrupt TCR signal strength despite unchanged affinity for TCR and can significantly impact the CD8+ T cell response to a viral escape mutant.

  12. Ampicillin/penicillin-binding protein interactions as a model drug-target system to optimize affinity pull-down and mass spectrometric strategies for target and pathway identification.

    Science.gov (United States)

    von Rechenberg, Moritz; Blake, Brian Kelly; Ho, Yew-Seng J; Zhen, Yuejun; Chepanoske, Cindy Lou; Richardson, Bonnie E; Xu, Nafei; Kery, Vladimir

    2005-05-01

    The identification and validation of the targets of active compounds identified in cell-based assays is an important step in preclinical drug development. New analytical approaches that combine drug affinity pull-down assays with mass spectrometry (MS) could lead to the identification of new targets and druggable pathways. In this work, we investigate a drug-target system consisting of ampicillin- and penicillin-binding proteins (PBPs) to evaluate and compare different amino-reactive resins for the immobilization of the affinity compound and mass spectrometric methods to identify proteins from drug affinity pull-down assays. First, ampicillin was immobilized onto various amino-reactive resins, which were compared in the ampicillin-PBP model with respect to their nonspecific binding of proteins from an Escherichia coli membrane extract. Dynal M-270 magnetic beads were chosen to further study the system as a model for capturing and identifying the targets of ampicillin, PBPs that were specifically and covalently bound to the immobilized ampicillin. The PBPs were identified, after in situ digestion of proteins bound to ampicillin directly on the beads, by using either one-dimensional (1-D) or two-dimensional (2-D) liquid chromatography (LC) separation techniques followed by tandem mass spectrometry (MS/MS) analysis. Alternatively, an elution with N-lauroylsarcosine (sarcosyl) from the ampicillin beads followed by in situ digestion and 2-D LC-MS/MS analysis identified proteins potentially interacting noncovalently with the PBPs or the ampicillin. The in situ approach required only little time, resources, and sample for the analysis. The combination of drug affinity pull-down assays with in situ digestion and 2-D LC-MS/MS analysis is a useful tool in obtaining complex information about a primary drug target as well as its protein interactors. PMID:15761956

  13. Two-dimensional Length Extraction of Ballistic Target from ISAR Images Using a New Scaling Method by Affine Registration

    Directory of Open Access Journals (Sweden)

    Jin Guanghu

    2014-09-01

    Full Text Available The length of ballistic target is one of the most important features for target recognition. It can be extracted from ISAR Images. Unlike from the optical image, the length extraction from ISAR image has two difficulties. The first one is that it is hard to get the actual position of scattering centres by the traditional target extraction method. The second one is that the ISAR image’s cross scale is not known because of the target’s complex rotation. Here we propose two methods to solve these problems. Firstly, we use clustering method to get scattering centers. Secondly we propose to get cross scale of the ISAR images by affine registration. Experiments verified that our approach is realisable and has good performance.Defence Science Journal, Vol. 64, No. 5, September 2014, pp.458-463, DOI:http://dx.doi.org/10.14429/dsj.64.5001

  14. High Affinity Antibodies against Influenza Characterize the Plasmablast Response in SLE Patients After Vaccination.

    Directory of Open Access Journals (Sweden)

    Kaval Kaur

    Full Text Available Breakdown of B cell tolerance is a cardinal feature of systemic lupus erythematosus (SLE. Increased numbers of autoreactive mature naïve B cells have been described in SLE patients and autoantibodies have been shown to arise from autoreactive and non-autoreactive precursors. How these defects, in the regulation of B cell tolerance and selection, influence germinal center (GC reactions that are directed towards foreign antigens has yet to be investigated. Here, we examined the characteristics of post-GC foreign antigen-specific B cells from SLE patients and healthy controls by analyzing monoclonal antibodies generated from plasmablasts induced specifically by influenza vaccination. We report that many of the SLE patients had anti-influenza antibodies with higher binding affinity and neutralization capacity than those from controls. Although overall frequencies of autoreactivity in the influenza-specific plasmablasts were similar for SLE patients and controls, the variable gene repertoire of influenza-specific plasmablasts from SLE patients was altered, with increased usage of JH6 and long heavy chain CDR3 segments. We found that high affinity anti-influenza antibodies generally characterize the plasmablast responses of SLE patients with low levels of autoreactivity; however, certain exceptions were noted. The high-avidity antibody responses in SLE patients may also be correlated with cytokines that are abnormally expressed in lupus. These findings provide insights into the effects of dysregulated immunity on the quality of antibody responses following influenza vaccination and further our understanding of the underlying abnormalities of lupus.

  15. High Affinity Antibodies against Influenza Characterize the Plasmablast Response in SLE Patients After Vaccination

    Science.gov (United States)

    Kaur, Kaval; Zheng, Nai-Ying; Smith, Kenneth; Huang, Min; Li, Lie; Pauli, Noel T.; Henry Dunand, Carole J.; Lee, Jane-Hwei; Morrissey, Michael; Wu, Yixuan; Joachims, Michelle L.; Munroe, Melissa E.; Lau, Denise; Qu, Xinyan; Krammer, Florian; Wrammert, Jens; Palese, Peter; Ahmed, Rafi; James, Judith A.; Wilson, Patrick C.

    2015-01-01

    Breakdown of B cell tolerance is a cardinal feature of systemic lupus erythematosus (SLE). Increased numbers of autoreactive mature naïve B cells have been described in SLE patients and autoantibodies have been shown to arise from autoreactive and non-autoreactive precursors. How these defects, in the regulation of B cell tolerance and selection, influence germinal center (GC) reactions that are directed towards foreign antigens has yet to be investigated. Here, we examined the characteristics of post-GC foreign antigen-specific B cells from SLE patients and healthy controls by analyzing monoclonal antibodies generated from plasmablasts induced specifically by influenza vaccination. We report that many of the SLE patients had anti-influenza antibodies with higher binding affinity and neutralization capacity than those from controls. Although overall frequencies of autoreactivity in the influenza-specific plasmablasts were similar for SLE patients and controls, the variable gene repertoire of influenza-specific plasmablasts from SLE patients was altered, with increased usage of JH6 and long heavy chain CDR3 segments. We found that high affinity anti-influenza antibodies generally characterize the plasmablast responses of SLE patients with low levels of autoreactivity; however, certain exceptions were noted. The high-avidity antibody responses in SLE patients may also be correlated with cytokines that are abnormally expressed in lupus. These findings provide insights into the effects of dysregulated immunity on the quality of antibody responses following influenza vaccination and further our understanding of the underlying abnormalities of lupus. PMID:25951191

  16. Affinity-Mediated Homogeneous Electrochemical Aptasensor on a Graphene Platform for Ultrasensitive Biomolecule Detection via Exonuclease-Assisted Target-Analog Recycling Amplification.

    Science.gov (United States)

    Ge, Lei; Wang, Wenxiao; Sun, Ximei; Hou, Ting; Li, Feng

    2016-02-16

    As is well-known, graphene shows a remarkable difference in affinity toward nonstructured single-stranded (ss) DNA and double-stranded (ds) DNA. This property makes it popular to prepare DNA-based optical sensors. In this work, taking this unique property of graphene in combination with the sensitive electrochemical transducer, we report a novel affinity-mediated homogeneous electrochemical aptasensor using graphene modified glassy carbon electrode (GCE) as the sensing platform. In this approach, the specific aptamer-target recognition is converted into an ultrasensitive electrochemical signal output with the aid of a novel T7 exonuclease (T7Exo)-assisted target-analog recycling amplification strategy, in which the ingeniously designed methylene blue (MB)-labeled hairpin DNA reporters are digested in the presence of target and, then, converted to numerous MB-labeled long ssDNAs. The distinct difference in differential pulse voltammetry response between the designed hairpin reporters and the generated long ssDNAs on the graphene/GCE allows ultrasensitive detection of target biomolecules. Herein, the design and working principle of this homogeneous electrochemical aptasensor were elucidated, and the working conditions were optimized. The gel electrophoresis results further demonstrate that the designed T7Exo-assisted target-analog recycling amplification strategy can work well. This electrochemical aptasensor realizes the detection of biomolecule in a homogeneous solution without immobilization of any bioprobe on electrode surface. Moreover, this versatile homogeneous electrochemical sensing system was used for the determination of biomolecules in real serum samples with satisfying results.

  17. T-cell receptor affinity and avidity defines antitumor response and autoimmunity in T-cell immunotherapy

    Science.gov (United States)

    Zhong, Shi; Malecek, Karolina; Johnson, Laura A.; Yu, Zhiya; Vega-Saenz de Miera, Eleazar; Darvishian, Farbod; McGary, Katelyn; Huang, Kevin; Boyer, Josh; Corse, Emily; Shao, Yongzhao; Rosenberg, Steven A.; Restifo, Nicholas P.; Osman, Iman; Krogsgaard, Michelle

    2013-01-01

    T cells expressing antigen-specific T-cell receptors (TCRs) can mediate effective tumor regression, but they often also are accompanied by autoimmune responses. To determine the TCR affinity threshold defining the optimal balance between effective antitumor activity and autoimmunity in vivo, we used a unique self-antigen system comprising seven human melanoma gp100(209–217)-specific TCRs spanning physiological affinities (1–100 μM). We found that in vitro and in vivo T-cell responses are determined by TCR affinity, except in one case that was compensated by substantial CD8 involvement. Strikingly, we found that T-cell antitumor activity and autoimmunity are closely coupled but plateau at a defined TCR affinity of 10 µM, likely due to diminished contribution of TCR affinity to avidity above the threshold. Together, these results suggest that a relatively low-affinity threshold is necessary for the immune system to avoid self-damage, given the close relationship between antitumor activity and autoimmunity. The low threshold, in turn, indicates that adoptive T-cell therapy treatment strategies using in vitro-generated high-affinity TCRs do not necessarily improve efficacy. PMID:23576742

  18. A high affinity kidney targeting by chitobionic acid-conjugated polysorbitol gene transporter alleviates unilateral ureteral obstruction in rats.

    Science.gov (United States)

    Islam, Mohammad Ariful; Kim, Sanghwa; Firdous, Jannatul; Lee, Ah-Young; Hong, Seong-Ho; Seo, Min Kyeong; Park, Tae-Eun; Yun, Cheol-Heui; Choi, Yun-Jaie; Chae, Chanhee; Cho, Chong-Su; Cho, Myung-Haing

    2016-09-01

    Aside from kidney transplantation - a procedure which is exceedingly dependent on donor-match and availability leading to excessive costs - there are currently no permanent treatments available which reverse kidney injury and failure. However, kidney-specific targeted gene therapy has outstanding potential to treat kidney-related dysfunction. Herein we report a novel kidney-specific targeted gene delivery system developed through the conjugation of chitobionic acid (CBA) to a polysorbitol gene transporter (PSGT) synthesized from sorbitol diacrylate and low molecular weight polyethylenimine (PEI) carrying hepatocyte growth factor (HGF) gene to alleviate unilateral ureteral obstruction (UUO) in rats. CBA-PSGT performed exceptionally well for targeted delivery of HGF to kidney tissues compared to its non-targeted counterparts (P type I and II), blood urea nitrogen (BUN), creatinine, and the expressions of ICAM-1, TIMP-1 and α-SMA which play a critical role in obstructive kidney functions. Therefore, CBA-PSGT should be further investigated because of its potential to alleviate UUO and kidney-related diseases using high affinity kidney targeting. PMID:27318934

  19. TCR affinity for thymoproteasome-dependent positively selecting peptides conditions antigen responsiveness in CD8(+) T cells.

    Science.gov (United States)

    Takada, Kensuke; Van Laethem, Francois; Xing, Yan; Akane, Kazuyuki; Suzuki, Haruhiko; Murata, Shigeo; Tanaka, Keiji; Jameson, Stephen C; Singer, Alfred; Takahama, Yousuke

    2015-10-01

    In the thymus, low-affinity T cell antigen receptor (TCR) engagement facilitates positive selection of a useful T cell repertoire. Here we report that TCR responsiveness of mature CD8(+) T cells is fine tuned by their affinity for positively selecting peptides in the thymus and that optimal TCR responsiveness requires positive selection on major histocompatibility complex class I-associated peptides produced by the thymoproteasome, which is specifically expressed in the thymic cortical epithelium. Thymoproteasome-independent positive selection of monoclonal CD8(+) T cells results in aberrant TCR responsiveness, homeostatic maintenance and immune responses to infection. These results demonstrate a novel aspect of positive selection, in which TCR affinity for positively selecting peptides produced by thymic epithelium determines the subsequent antigen responsiveness of mature CD8(+) T cells in the periphery.

  20. α4βδ GABA(A) receptors are high-affinity targets for γ-hydroxybutyric acid (GHB).

    Science.gov (United States)

    Absalom, Nathan; Eghorn, Laura F; Villumsen, Inge S; Karim, Nasiara; Bay, Tina; Olsen, Jesper V; Knudsen, Gitte M; Bräuner-Osborne, Hans; Frølund, Bente; Clausen, Rasmus P; Chebib, Mary; Wellendorph, Petrine

    2012-08-14

    γ-Hydroxybutyric acid (GHB) binding to brain-specific high-affinity sites is well-established and proposed to explain both physiological and pharmacological actions. However, the mechanistic links between these lines of data are unknown. To identify molecular targets for specific GHB high-affinity binding, we undertook photolinking studies combined with proteomic analyses and identified several GABA(A) receptor subunits as possible candidates. A subsequent functional screening of various recombinant GABA(A) receptors in Xenopus laevis oocytes using the two-electrode voltage clamp technique showed GHB to be a partial agonist at αβδ- but not αβγ-receptors, proving that the δ-subunit is essential for potency and efficacy. GHB showed preference for α4 over α(1,2,6)-subunits and preferably activated α4β1δ (EC(50) = 140 nM) over α4β(2/3)δ (EC(50) = 8.41/1.03 mM). Introduction of a mutation, α4F71L, in α4β1(δ)-receptors completely abolished GHB but not GABA function, indicating nonidentical binding sites. Radioligand binding studies using the specific GHB radioligand [(3)H](E,RS)-(6,7,8,9-tetrahydro-5-hydroxy-5H-benzocyclohept-6-ylidene)acetic acid showed a 39% reduction (P = 0.0056) in the number of binding sites in α4 KO brain tissue compared with WT controls, corroborating the direct involvement of the α4-subunit in high-affinity GHB binding. Our data link specific GHB forebrain binding sites with α4-containing GABA(A) receptors and postulate a role for extrasynaptic α4δ-containing GABA(A) receptors in GHB pharmacology and physiology. This finding will aid in elucidating the molecular mechanisms behind the proposed function of GHB as a neurotransmitter and its unique therapeutic effects in narcolepsy and alcoholism.

  1. Affinity peptide developed by phage display selection for targeting gastric cancer

    Institute of Scientific and Technical Information of China (English)

    Wen-Jie Zhang; Yan-Xia Sui; Arun Budha; Jian-Bao Zheng; Xue-Jun Sun; Ying-Chun Hou; Thomas D Wang; Shao-Ying Lu

    2012-01-01

    AIM:To develop an affinity peptide that binds to gastric cancer used for the detection of early gastric cancer.METHODS:A peptide screen was performed by biopanning the PhD-12 phage display library,clearing non-specific binders against tumor-adjacent normal appearing gastric mucosa and obtaining selective binding against freshly harvested gastric cancer tissues.Tumortargeted binding of selected peptides was confirmed by bound phage counts,enzyme-linked immunosorbent assay,competitive inhibition,fluorescence microscopy and semi-quantitative analysis on immunohistochemistry using different types of cancer tissues.RESULTS:Approximately 92.8% of the non-specific phage clones were subtracted from the original phage library after two rounds of biopanning against normalappearing gastric mucosa.After the third round of positive screening,the peptide sequence AADNAKTKSFPV (AAD) appeared in 25% (12/48) of the analyzed phages.For the control peptide,these values were 6.8 ± 2.3,5.1 ± 1.7,3.5 ± 2.1,4.6 ± 1.9 and 1.1 ± 0.5,respectively.The values for AAD peptide were statistically significant (P < 0.01) for gastric cancer as compared with other histological classifications and control peptide.CONCLUSION:A novel peptide is discovered to have a specific binding activity to gastric cancer,and can be used to distinguish neoplastic from normal gastric mucosa,demonstrating the potential for early cancer detection on endoscopy.

  2. α4βδ GABA receptors are high-affinity targets for γ-hydroxybutyric acid (GHB)

    DEFF Research Database (Denmark)

    Absalom, N.; Karim, N.; Eghorn, L.F.;

    2012-01-01

    γ-Hydroxybutyric acid (GHB) binding to brain-specific high-affinity sites is well-established and proposed to explain both physiological and pharmacological actions. However, the mechanistic links between these lines of data are unknown. To identify molecular targets for specific GHB high......-affinity binding, we undertook photolinking studies combined with proteomic analyses and identified several GABA receptor subunits as possible candidates. A subsequent functional screening of various recombinant GABA receptors in Xenopus laevis oocytes using the two-electrode voltage clamp technique showed GHB...... to be a partial agonist at αβδ- but not αβγ-receptors, proving that the δ-subunit is essential for potency and efficacy. GHB showed preference for α4 over α(1,2,6)-subunits and preferably activated α4β1δ (EC = 140 nM) over α4β(2/3)δ (EC = 8.41/1.03 mM). Introduction of a mutation, α4F71L, in α4β 1(δ...

  3. Studies on lipase-affinity adsorption using response-surface analysis.

    Science.gov (United States)

    Kamimura, E S; Medieta, O; Rodrigues, M I; Maugeri, F

    2001-06-01

    Lipases are widely distributed enzymes that can be obtained from animals, plants and micro-organisms. Coupling lipases with a wide range of substrates allows the opportunity for synthesis of optically pure pharmaceutical preparations, flavour compounds and other food additives. Affinity chromatography owes its power as a purification method to specific biological interactions. Response-surface analysis was chosen to study column efficiency. This method allows the understanding of interactions between variables with advantages over conventional methods, which involve changing one variable while fixing others at certain levels. The aim of this work was to study the influence of the ratio bed height/column diameter (L/D) and superficial velocity (V) on the column efficiency. The experimental design involved the two variables, L/D (2-10) and v (1-2 cm/min), at five levels. Lipase was obtained from Geotrichum sp. culture in a complex medium composed of 5% corn-steep liquor, 0.5% NH(4)NO(3) and 1% olive oil at 30 degrees C, with 1VVM (air volume/medium volume per min) aeration and 400 rev./min agitation. Maximum lipase activity was 19 units/ml after almost 9 h of fermentation. This lipase could potentially be used in esterification reactions to increase the content of gamma-linolenic acid and to produce bioaromas for food industries. The adsorption assays were carried out in a fixed-bed column with an affinity adsorbent, which was obtained by reaction of a gel with oleic acid as ligand. Breakthrough curves were obtained for all experiments. It has been shown that the lower the values of both L/D and v, the higher the column efficiency (maximum 65.43%). Also, it was observed from the response surface that the efficiency reached a minimum at an L/D of around 8.

  4. An In-tether Chiral Center Modulates the Helicity, Cell Permeability, and Target Binding Affinity of a Peptide.

    Science.gov (United States)

    Hu, Kuan; Geng, Hao; Zhang, Qingzhou; Liu, Qisong; Xie, Mingsheng; Sun, Chengjie; Li, Wenjun; Lin, Huacan; Jiang, Fan; Wang, Tao; Wu, Yun-Dong; Li, Zigang

    2016-07-01

    The addition of a precisely positioned chiral center in the tether of a constrained peptide is reported, yielding two separable peptide diastereomers with significantly different helicity, as supported by circular dichroism (CD) and NMR spectroscopy. Single crystal X-ray diffraction analysis suggests that the absolute configuration of the in-tether chiral center in helical form is R, which is in agreement with theoretical simulations. The relationship between the secondary structure of the short peptides and their biochemical/biophysical properties remains elusive, largely because of the lack of proper controls. The present strategy provides the only method for investigating the influence of solely conformational differences upon the biochemical/biophysical properties of peptides. The significant differences in permeability and target binding affinity between the peptide diastereomers demonstrate the importance of helical conformation. PMID:27167181

  5. Functional analysis and binding affinity of tomato ethylene response factors provide insight on the molecular bases of plant differential responses to ethylene

    Directory of Open Access Journals (Sweden)

    Pirrello Julien

    2012-10-01

    Full Text Available Abstract Background The phytohormone ethylene is involved in a wide range of developmental processes and in mediating plant responses to biotic and abiotic stresses. Ethylene signalling acts via a linear transduction pathway leading to the activation of Ethylene Response Factor genes (ERF which represent one of the largest gene families of plant transcription factors. How an apparently simple signalling pathway can account for the complex and widely diverse plant responses to ethylene remains yet an unanswered question. Building on the recent release of the complete tomato genome sequence, the present study aims at gaining better insight on distinctive features among ERF proteins. Results A set of 28 cDNA clones encoding ERFs in the tomato (Solanum lycopersicon were isolated and shown to fall into nine distinct subclasses characterised by specific conserved motifs most of which with unknown function. In addition of being able to regulate the transcriptional activity of GCC-box containing promoters, tomato ERFs are also shown to be active on promoters lacking this canonical ethylene-responsive-element. Moreover, the data reveal that ERF affinity to the GCC-box depends on the nucleotide environment surrounding this cis-acting element. Site-directed mutagenesis revealed that the nature of the flanking nucleotides can either enhance or reduce the binding affinity, thus conferring the binding specificity of various ERFs to target promoters. Based on their expression pattern, ERF genes can be clustered in two main clades given their preferential expression in reproductive or vegetative tissues. The regulation of several tomato ERF genes by both ethylene and auxin, suggests their potential contribution to the convergence mechanism between the signalling pathways of the two hormones. Conclusions The data reveal that regions flanking the core GCC-box sequence are part of the discrimination mechanism by which ERFs selectively bind to their target

  6. Structure-based rational design of a Toll-like receptor 4 (TLR4 decoy receptor with high binding affinity for a target protein.

    Directory of Open Access Journals (Sweden)

    Jieun Han

    Full Text Available Repeat proteins are increasingly attracting much attention as alternative scaffolds to immunoglobulin antibodies due to their unique structural features. Nonetheless, engineering interaction interface and understanding molecular basis for affinity maturation of repeat proteins still remain a challenge. Here, we present a structure-based rational design of a repeat protein with high binding affinity for a target protein. As a model repeat protein, a Toll-like receptor4 (TLR4 decoy receptor composed of leucine-rich repeat (LRR modules was used, and its interaction interface was rationally engineered to increase the binding affinity for myeloid differentiation protein 2 (MD2. Based on the complex crystal structure of the decoy receptor with MD2, we first designed single amino acid substitutions in the decoy receptor, and obtained three variants showing a binding affinity (K(D one-order of magnitude higher than the wild-type decoy receptor. The interacting modes and contributions of individual residues were elucidated by analyzing the crystal structures of the single variants. To further increase the binding affinity, single positive mutations were combined, and two double mutants were shown to have about 3000- and 565-fold higher binding affinities than the wild-type decoy receptor. Molecular dynamics simulations and energetic analysis indicate that an additive effect by two mutations occurring at nearby modules was the major contributor to the remarkable increase in the binding affinities.

  7. Targeted therapy against human lung cancer in nude mice by high-affinity recombinant antimesothelin single-chain Fv immunotoxin.

    Science.gov (United States)

    Fan, Dominic; Yano, Seiji; Shinohara, Hisashi; Solorzano, Carmen; Van Arsdall, Melissa; Bucana, Corazon D; Pathak, Sen; Kruzel, Ewa; Herbst, Roy S; Onn, Amir; Roach, Jennifer S; Onda, Masanori; Wang, Qing-cheng; Pastan, Ira; Fidler, Isaiah J

    2002-06-01

    Several tumors, including mesothelioma and ovarian cancer, can overexpress mesothelin, a glycosylphosphatidylinositol-linked differentiation glycoprotein. The membrane-bound type of mesothelin is found in the blood of cancer patients at a very low level, which makes mesothelin a good candidate for targeted therapy of certain cancers. An antimesothelin disulfide-linked Fv (SS1 Fv) was fused to a truncated mutant of Pseudomonas exotoxin A to produce the recombinant immunotoxin SS1(dsFv)-PE38, which has a high binding affinity to mesothelin (Kd = 0.7 nM). Our studies in vitro showed that SS1(dsFv)-PE38 is significantly more cytotoxic to the high-mesothelin-producing NCI-H226 human non-small cell lung cancer cells than to human lung adenocarcinoma PC14PE6 cells, which do not express mesothelin. When administered at a nontoxic dose of 500 microg/kg on days 7, 9, and 11 to nude mice injected i.v. with the two human lung cancer cell lines, SS1(dsFv)-PE38 selectively inhibited experimental lung metastases produced by the mesothelin-producing NCI-H226 cells. Our data indicate that mesothelin-producing squamous cell carcinoma of the lung may be a good target for this immunotoxin. PMID:12479219

  8. Pegylated Trastuzumab Fragments Acquire an Increased in Vivo Stability but Show a Largely Reduced Affinity for the Target Antigen.

    Science.gov (United States)

    Selis, Fabio; Focà, Giuseppina; Sandomenico, Annamaria; Marra, Carla; Di Mauro, Concetta; Saccani Jotti, Gloria; Scaramuzza, Silvia; Politano, Annalisa; Sanna, Riccardo; Ruvo, Menotti; Tonon, Giancarlo

    2016-01-01

    PEGylation of biomolecules is a major approach to increase blood stream half-life, stability and solubility of biotherapeutics and to reduce their immunogenicity, aggregation potential and unspecific interactions with other proteins and tissues. Antibodies have generally long half-lives due to high molecular mass and stability toward proteases, however their size lowers to some extent their potential because of a reduced ability to penetrate tissues, especially those of tumor origin. Fab or otherwise engineered smaller fragments are an alternative but are less stable and are much less well retained in circulation. We have here investigated the effects of various PEGylations on the binding properties and in vivo half-life of Fab fragments derived from the enzymatic splitting of Trastuzumab. We find that PEGylation increases the half-life of the molecules but also strongly affects the ability to recognize the target antigen in a way that is dependent on the extent and position of the chemical modification. Data thus support the concept that polyethylene glycol (PEG) conjugation on Trastuzumab Fabs increases half-life but reduces their affinity and this is a fine balance, which must be carefully considered for the design of strategies based on the use of antibody fragments.

  9. Dissecting the Binding Mode of Low Affinity Phage Display Peptide Ligands to Protein Targets by Hydrogen/Deuterium Exchange Coupled to Mass Spectrometry

    DEFF Research Database (Denmark)

    Leurs, Ulrike; Lohse, Brian; Ming, Shonoi A;

    2014-01-01

    of hydrogen/deuterium exchange mass spectrometry (HDX-MS) to characterize interactions of low affinity peptides with their cognate protein targets. The HDX-MS workflow was optimized to accurately detect low-affinity peptide-protein interactions by use of ion mobility, electron transfer dissociation, non...... of KDM4C, indicating distinct binding modes. In contrast, the perturbation site of another PD-selected peptide inhibiting the function of KDM1A maps to a GST-tag. Our results demonstrate that HDX-MS can validate and map weak peptide-protein interactions, and pave the way for understanding and optimizing...

  10. Kinetic Characterization of a Panel of High-Affinity Monoclonal Antibodies Targeting Ricin and Recombinant Re-Formatting for Biosensor Applications

    Directory of Open Access Journals (Sweden)

    Michelle Cummins

    2014-05-01

    Full Text Available Ricin is a potent glycoprotein toxin that is structurally composed of two subunits joined via a disulfide bond: a ~30 kDa subunit A (RTA and a ~32 kDa subunit B (RTB. There are fears of ricin being used as a weapon for warfare and terrorism and, as such, there is an increasing need for the development of immunodiagnostic reagents targeted towards this toxin. This article describes the production and characterization of a panel of six ricin-specific monoclonal IgG antibodies (mAbs, previously selected based upon their ability to inhibit ricin-mediated killing of cultured cells. Subsequent epitope binding analysis using the surface plasmon resonance (SPR array biosensor (ProteOn XPR36 indicated three distinct, non-competitive binding epitopes (“bins”. The association (ka and dissociation (kd rate constants and binding affinities (KD of each of the mAbs to ricin were also determined by SPR using Biacore T100 instrument. Affinities (KD ranged from 0.1 nM to 9 nM. We present the coding sequences of the variable domains of the six mAbs, the expression, kinetic and cytotoxicity assays for two recombinant Fab (rFab fragments and demonstrate a rFab affinity improvement by chain-shuffling. Together, these antibodies and constituent rFabs represent a panel of reagents for high-affinity recognition of ricin with potential national security biosensor applications.

  11. Affinity Maturation of Monoclonal Antibody 1E11 by Targeted Randomization in CDR3 Regions Optimizes Therapeutic Antibody Targeting of HER2-Positive Gastric Cancer.

    Science.gov (United States)

    Ko, Bong-Kook; Choi, Soyoung; Cui, Lei Guang; Lee, Young-Ha; Hwang, In-Sik; Kim, Kyu-Tae; Shim, Hyunbo; Lee, Jong-Seo

    2015-01-01

    Anti-HER2 murine monoclonal antibody 1E11 has strong and synergistic anti-tumor activity in HER2-overexpressing gastric cancer cells when used in combination with trastuzumab. We presently optimized this antibody for human therapeutics. First, the complementarity determining regions (CDRs) of the murine antibody were grafted onto human germline immunoglobulin variable genes. No difference in affinity and biological activity was observed between chimeric 1E11 (ch1E11) and humanized 1E11 (hz1E11). Next, affinity maturation of hz1E11 was performed by the randomization of CDR-L3 and H3 residues followed by stringent biopanning selection. Milder selection pressure favored the selection of more diverse clones, whereas higher selection stringency resulted in the convergence of the panning output to a smaller number of clones with improved affinity. Clone 1A12 had four amino acid substitutions in CDR-L3, and showed a 10-fold increase in affinity compared to the parental clone and increased potency in an in vitro anti-proliferative activity assay with HER2-overepxressing gastric cancer cells. Clone 1A12 inhibited tumor growth of NCI-N87 xenograft model with similar efficacy to trastuzumab alone, and the combination treatment of 1A12 and trastuzumab completely removed the established tumors. These results suggest that humanized and affinity matured monoclonal antibody 1A12 is a highly optimized molecule for future therapeutic development against HER2-positive tumors.

  12. A dualistic conformational response to substrate binding in the human serotonin transporter reveals a high affinity state for serotonin

    DEFF Research Database (Denmark)

    Bjerregaard, Henriette; Severinsen, Kasper; Said, Saida;

    2015-01-01

    Serotonergic neurotransmission is modulated by the membrane-embedded serotonin transporter (SERT). SERT mediates the reuptake of serotonin into the presynaptic neurons. Conformational changes in SERT occur upon binding of ions and substrate and are crucial for translocation of serotonin across...... that were sensitized to detect a more outward-facing conformation of SERT. We found a novel high affinity outward-facing conformational state of the human SERT induced by serotonin. The ionic requirements for this new conformational response to serotonin mirror the ionic requirements for translocation...

  13. The tumor targeted superantigen ABR-217620 selectively engages TRBV7-9 and exploits TCR-pMHC affinity mimicry in mediating T cell cytotoxicity.

    Directory of Open Access Journals (Sweden)

    Gunnar Hedlund

    Full Text Available The T lymphocytes are the most important effector cells in immunotherapy of cancer. The conceptual objective for developing the tumor targeted superantigen (TTS ABR-217620 (naptumomab estafenatox, 5T4Fab-SEA/E-120, now in phase 3 studies for advanced renal cell cancer, was to selectively coat tumor cells with cytotoxic T lymphocytes (CTL target structures functionally similar to natural CTL pMHC target molecules. Here we present data showing that the molecular basis for the anti-tumor activity by ABR-217620 resides in the distinct interaction between the T cell receptor β variable (TRBV 7-9 and the engineered superantigen (Sag SEA/E-120 in the fusion protein bound to the 5T4 antigen on tumor cells. Multimeric but not monomeric ABR-217620 selectively stains TRBV7-9 expressing T lymphocytes from human peripheral blood similar to antigen specific staining of T cells with pMHC tetramers. SEA/E-120 selectively activates TRBV7-9 expressing T lymphocytes resulting in expansion of the subset. ABR-217620 selectively triggers TRBV7-9 expressing cytotoxic T lymphocytes to kill 5T4 positive tumor cells. Furthermore, ABR-217620 activates TRBV7-9 expressing T cell line cells in the presence of cell- and bead-bound 5T4 tumor antigen. Surface plasmon resonance analysis revealed that ABR-217620 binds to 5T4 with high affinity, to TRBV7-9 with low affinity and to MHC class II with very low affinity. The T lymphocyte engagement by ABR-217620 is constituted by displaying high affinity binding to the tumor cells (KD approximately 1 nM and with the mimicry of natural productive immune TCR-pMHC contact using affinities of around 1 µM. This difference in kinetics between the two components of the ABR-217620 fusion protein will bias the binding towards the 5T4 target antigen, efficiently activating T-cells via SEA/E-120 only when presented by the tumor cells.

  14. Consumer responses to ethnic targeted marketing

    NARCIS (Netherlands)

    A.-S.I.A. Lenoir (Anne-Sophie); S. Puntoni (Stefano)

    2014-01-01

    markdownabstract__Abstract__ Marketing is impacted more than ever by demographic change, to the extent that practitioners targeting ethnic groups should re-think their approach depending upon the strength with which different generations identify with their cultural heritage.

  15. Design and Synthesis of High-Affinity Dimeric Inhibitors Targeting the Interactions between Gephyrin and Inhibitory Neurotransmitter Receptors

    DEFF Research Database (Denmark)

    Maric, Hans-Michael; Kasaragod, Vikram Babu; Kedström, Linda Maria Haugaard;

    2015-01-01

    Gephyrin is the central scaffolding protein for inhibitory neurotransmitter receptors in the brain. Here we describe the development of dimeric peptides that inhibit the interaction between gephyrin and these receptors, a process which is fundamental to numerous synaptic functions and diseases...... of the brain. We first identified receptor-derived minimal gephyrin-binding peptides that displayed exclusive binding towards native gephyrin from brain lysates. We then designed and synthesized a series of dimeric ligands, which led to a remarkable 1220-fold enhancement of the gephyrin affinity (KD =6.8 n......M). In X-ray crystal structures we visualized the simultaneous dimer-to-dimer binding in atomic detail, revealing compound-specific binding modes. Thus, we defined the molecular basis of the affinity-enhancing effect of multivalent gephyrin inhibitors and provide conceptually novel compounds...

  16. Proteochemometric modelling coupled to in silico target prediction: an integrated approach for the simultaneous prediction of polypharmacology and binding affinity/potency of small molecules.

    Science.gov (United States)

    Paricharak, Shardul; Cortés-Ciriano, Isidro; IJzerman, Adriaan P; Malliavin, Thérèse E; Bender, Andreas

    2015-01-01

    The rampant increase of public bioactivity databases has fostered the development of computational chemogenomics methodologies to evaluate potential ligand-target interactions (polypharmacology) both in a qualitative and quantitative way. Bayesian target prediction algorithms predict the probability of an interaction between a compound and a panel of targets, thus assessing compound polypharmacology qualitatively, whereas structure-activity relationship techniques are able to provide quantitative bioactivity predictions. We propose an integrated drug discovery pipeline combining in silico target prediction and proteochemometric modelling (PCM) for the respective prediction of compound polypharmacology and potency/affinity. The proposed pipeline was evaluated on the retrospective discovery of Plasmodium falciparum DHFR inhibitors. The qualitative in silico target prediction model comprised 553,084 ligand-target associations (a total of 262,174 compounds), covering 3,481 protein targets and used protein domain annotations to extrapolate predictions across species. The prediction of bioactivities for plasmodial DHFR led to a recall value of 79% and a precision of 100%, where the latter high value arises from the structural similarity of plasmodial DHFR inhibitors and T. gondii DHFR inhibitors in the training set. Quantitative PCM models were then trained on a dataset comprising 20 eukaryotic, protozoan and bacterial DHFR sequences, and 1,505 distinct compounds (in total 3,099 data points). The most predictive PCM model exhibited R (2) 0 test and RMSEtest values of 0.79 and 0.59 pIC50 units respectively, which was shown to outperform models based exclusively on compound (R (2) 0 test/RMSEtest = 0.63/0.78) and target information (R (2) 0 test/RMSEtest = 0.09/1.22), as well as inductive transfer knowledge between targets, with respective R (2) 0 test and RMSEtest values of 0.76 and 0.63 pIC50 units. Finally, both methods were integrated to predict the protein

  17. Stimulus-responsive nanopreparations for tumor targeting.

    Science.gov (United States)

    Zhu, Lin; Torchilin, Vladimir P

    2013-01-01

    Nanopreparations such as liposomes, micelles, polymeric and inorganic nanoparticles, and small molecule/nucleic acid/protein conjugates have demonstrated various advantages over "naked" therapeutic molecules. These nanopreparations can be further engineered with functional moieties to improve their performance in terms of circulation longevity, targetability, enhanced intracellular penetration, carrier-mediated enhanced visualization, and stimuli-sensitivity. The idea of application of a stimulus-sensitive drug or imaging agent delivery system for tumor targeting is based on the significant abnormalities in the tumor microenvironment and its cells, such as an acidic pH, altered redox potential, up-regulated proteins and hyperthermia. These internal conditions as well as external stimuli, such as magnetic field, ultrasound and light, can be used to modify the behavior of the nanopreparations that control drug release, improve drug internalization, control the intracellular drug fate and even allow for certain physical interactions, resulting in an enhanced tumor targeting and antitumor effect. This article provides a critical view of current stimulus-sensitive drug delivery strategies and possible future directions in tumor targeting with primary focus on the combined use of stimulus-sensitivity with other strategies in the same nanopreparation, including multifunctional nanopreparations and theranostics. PMID:22869005

  18. Insights from the Fungus Fusarium oxysporum Point to High Affinity Glucose Transporters as Targets for Enhancing Ethanol Production from Lignocellulose

    Science.gov (United States)

    Ali, Shahin S.; Nugent, Brian; Mullins, Ewen; Doohan, Fiona M.

    2013-01-01

    Ethanol is the most-widely used biofuel in the world today. Lignocellulosic plant biomass derived from agricultural residue can be converted to ethanol via microbial bioprocessing. Fungi such as Fusarium oxysporum can simultaneously saccharify straw to sugars and ferment sugars to ethanol. But there are many bottlenecks that need to be overcome to increase the efficacy of microbial production of ethanol from straw, not least enhancement of the rate of fermentation of both hexose and pentose sugars. This research tested the hypothesis that the rate of sugar uptake by F. oxysporum would enhance the ethanol yields from lignocellulosic straw and that high affinity glucose transporters can enhance ethanol yields from this substrate. We characterized a novel hexose transporter (Hxt) from this fungus. The F. oxysporum Hxt represents a novel transporter with homology to yeast glucose signaling/transporter proteins Rgt2 and Snf3, but it lacks their C-terminal domain which is necessary for glucose signalling. Its expression level decreased with increasing glucose concentration in the medium and in a glucose uptake study the Km(glucose) was 0.9 mM, which indicated that the protein is a high affinity glucose transporter. Post-translational gene silencing or over expression of the Hxt in F. oxysporum directly affected the glucose and xylose transport capacity and ethanol yielded by F. oxysporum from straw, glucose and xylose. Thus we conclude that this Hxt has the capacity to transport both C5 and C6 sugars and to enhance ethanol yields from lignocellulosic material. This study has confirmed that high affinity glucose transporters are ideal candidates for improving ethanol yields from lignocellulose because their activity and level of expression is high in low glucose concentrations, which is very common during the process of consolidated processing. PMID:23382943

  19. Targeted drug induces responses in aggressive lymphomas

    Science.gov (United States)

    Preliminary results from clinical trials in a subtype of lymphoma show that for a number of patients whose disease was not cured by other treatments, the drug ibrutinib can provide significant anti-cancer responses with modest side effects.

  20. Evidence of self-affine multiplicity fluctuation of target residues in 84Kr-AgBr interactions at 1.7 AGeV

    Institute of Scientific and Technical Information of China (English)

    ZHANG Dong-Hai; LI Hui-Ling

    2009-01-01

    Self-afline multiplicity scaling is investigated in the framework of a two-dimensional factorial mo-ment methodology using the concept of the Hurst exponent (H). Analyzing the experimental data of target evaporated fragments emitted in84Kr-AgBr interactions at 1.7 AGeV revealed that the best power law behav-ior is exhibited for H = 0.3 indicating a self-affine multiplicity fluctuation pattern. A signal of multifractality is also observed from knowledge of the anomalous fractal dimension dq extracted from the intermittency exponent aq of the anisotropic phase space scenario.

  1. Stimulus-responsive nanopreparations for tumor targeting

    OpenAIRE

    ZHU, LIN; Torchilin, Vladimir P.

    2013-01-01

    Nanopreparations such as liposomes, micelles, polymeric and inorganic nanoparticles, and small molecule/nucleic acid/protein conjugates have demonstrated various advantages versus “naked” therapeutic molecules. These nanopreparations can be further engineered with functional moieties to improve their performance in terms of circulation longevity, targetability, enhanced intracellular penetration; carrier-mediated enhanced visualization, and stimuli-sensitivity. The idea of application of a st...

  2. Integration of Affinity Selection-Mass Spectrometry and Functional Cell-Based Assays to Rapidly Triage Druggable Target Space within the NF-κB Pathway.

    Science.gov (United States)

    Kutilek, Victoria D; Andrews, Christine L; Richards, Matthew P; Xu, Zangwei; Sun, Tianxiao; Chen, Yiping; Hashke, Andrew; Smotrov, Nadya; Fernandez, Rafael; Nickbarg, Elliott B; Chamberlin, Chad; Sauvagnat, Berengere; Curran, Patrick J; Boinay, Ryan; Saradjian, Peter; Allen, Samantha J; Byrne, Noel; Elsen, Nathaniel L; Ford, Rachael E; Hall, Dawn L; Kornienko, Maria; Rickert, Keith W; Sharma, Sujata; Shipman, Jennifer M; Lumb, Kevin J; Coleman, Kevin; Dandliker, Peter J; Kariv, Ilona; Beutel, Bruce

    2016-07-01

    The primary objective of early drug discovery is to associate druggable target space with a desired phenotype. The inability to efficiently associate these often leads to failure early in the drug discovery process. In this proof-of-concept study, the most tractable starting points for drug discovery within the NF-κB pathway model system were identified by integrating affinity selection-mass spectrometry (AS-MS) with functional cellular assays. The AS-MS platform Automated Ligand Identification System (ALIS) was used to rapidly screen 15 NF-κB proteins in parallel against large-compound libraries. ALIS identified 382 target-selective compounds binding to 14 of the 15 proteins. Without any chemical optimization, 22 of the 382 target-selective compounds exhibited a cellular phenotype consistent with the respective target associated in ALIS. Further studies on structurally related compounds distinguished two chemical series that exhibited a preliminary structure-activity relationship and confirmed target-driven cellular activity to NF-κB1/p105 and TRAF5, respectively. These two series represent new drug discovery opportunities for chemical optimization. The results described herein demonstrate the power of combining ALIS with cell functional assays in a high-throughput, target-based approach to determine the most tractable drug discovery opportunities within a pathway. PMID:26969322

  3. Identification of Thioredoxin Disulfide Targets Using a Quantitative Proteomics Approach Based on Isotope-Coded Affinity Tags

    DEFF Research Database (Denmark)

    Hägglund, Per; Bunkenborg, Jakob; Maeda, Kenji;

    2008-01-01

    , protein extract of embryos from germinated barley seeds was treated +/- Trx, and thiols released from target protein disulfides were irreversibly blocked with iodoacetamide. The remaining cysteine residues in the Trx-treated and the control (-Trx) samples were then chemically reduced and labeled...

  4. Affinity maturation of a novel antagonistic human monoclonal antibody with a long VH CDR3 targeting the Class A GPCR formyl-peptide receptor 1.

    Science.gov (United States)

    Douthwaite, Julie A; Sridharan, Sudharsan; Huntington, Catherine; Hammersley, Jayne; Marwood, Rose; Hakulinen, Jonna K; Ek, Margareta; Sjögren, Tove; Rider, David; Privezentzev, Cyril; Seaman, Jonathan C; Cariuk, Peter; Knights, Vikki; Young, Joyce; Wilkinson, Trevor; Sleeman, Matthew; Finch, Donna K; Lowe, David C; Vaughan, Tristan J

    2015-01-01

    Therapeutic monoclonal antibodies targeting G-protein-coupled receptors (GPCRs) are desirable for intervention in a wide range of disease processes. The discovery of such antibodies is challenging due to a lack of stability of many GPCRs as purified proteins. We describe here the generation of Fpro0165, a human anti-formyl peptide receptor 1 (FPR1) antibody generated by variable domain engineering of an antibody derived by immunization of transgenic mice expressing human variable region genes. Antibody isolation and subsequent engineering of affinity, potency and species cross-reactivity using phage display were achieved using FPR1 expressed on HEK cells for immunization and selection, along with calcium release cellular assays for antibody screening. Fpro0165 shows full neutralization of formyl peptide-mediated activation of primary human neutrophils. A crystal structure of the Fpro0165 Fab shows a long, protruding VH CDR3 of 24 amino acids and in silico docking with a homology model of FPR1 suggests that this long VH CDR3 is critical to the predicted binding mode of the antibody. Antibody mutation studies identify the apex of the long VH CDR3 as key to mediating the species cross-reactivity profile of the antibody. This study illustrates an approach for antibody discovery and affinity engineering to typically intractable membrane proteins such as GPCRs.

  5. Inverse Thermal-Responsive Glyco-Polypeptide Polymer for One-Pot Glyco-Affinity Proteomic

    Institute of Scientific and Technical Information of China (English)

    Xue-Long Sun

    2005-01-01

    @@ 1Introduction Proteins are ultimately responsible for the biological processes in cells, body fluids, and tissue specimens.This presents enormous challenges to the field of proteomics, which aims to identify, characterize and assign biological functions of all proteins. Determining individual protein in complex biological samples often requires some type of separation as a prerequisite for its measurement. The complexities of chemical structure and in the physiological function of every protein contribute to the problems encountered when trying to separate these biomolecules. A number of techniques exist for the separation of proteins, however, a separation technique that satisfies speed of analysis, selectivity, sensitivity, adequate throughput capacity, and affordable cost of analysis is unavailable yet.

  6. A high-affinity [18 F]-labeled phosphoramidate peptidomimetic PSMA-targeted inhibitor for PET imaging of prostate cancer

    International Nuclear Information System (INIS)

    Introduction: In this study, a structurally modified phosphoramidate scaffold, with improved prostate-specific membrane antigen (PSMA) avidity, stability and in vivo characteristics, as a PET imaging agent for prostate cancer (PCa), was prepared and evaluated. Methods: p-Fluorobenzoyl-aminohexanoate and 2-(3-hydroxypropyl)glycine were introduced into the PSMA-targeting scaffold yielding phosphoramidate 5. X-ray crystallography was performed on the PSMA/5 complex. [18F]5 was synthesized, and cell uptake and internalization studies were conducted in PSMA(+) LNCaP and CWR22Rv1 cells and PSMA(−) PC-3 cells. In vivo PET imaging and biodistribution studies were performed at 1 and 4 h post injection in mice bearing CWR22Rv1 tumor, with or without blocking agent. Results: The crystallographic data showed interaction of the p-fluorobenzoyl group with an arene-binding cleft on the PSMA surface. In vitro studies revealed elevated uptake of [18F]5 in PSMA(+) cells (2.2% in CWR22Rv1 and 12.1% in LNCaP) compared to PSMA(−) cells (0.08%) at 4 h. In vivo tumor uptake of 2.33% ID/g and tumor-to-blood ratio of 265:1 was observed at 4 h. Conclusions: We have successfully synthesized, radiolabeled and evaluated a new PSMA-targeted PET agent. The crystal structure of the PSMA/5 complex highlighted the interactions within the arene-binding cleft contributing to the overall complex stability. The high target uptake and rapid non-target clearance exhibited by [18F]5 in PSMA(+) xenografts substantiates its potential use for PET imaging of PCa. Advances in Knowledge: The only FDA-approved imaging agent for PCa, Prostascint®, targets PSMA but suffers from inherent shortcomings. The data acquired in this manuscript confirmed that our new generation of [18F]-labeled PSMA inhibitor exhibited promising in vivo performance as a PET imaging agent for PCa and is well-positioned for subsequent clinical trials. Implications for Patient Care Our preliminary data demonstrate that this tracer

  7. XGFR*, a novel affinity-matured bispecific antibody targeting IGF-1R and EGFR with combined signaling inhibition and enhanced immune activation for the treatment of pancreatic cancer.

    Science.gov (United States)

    Schanzer, Juergen M; Wartha, Katharina; Moessner, Ekkehard; Hosse, Ralf J; Moser, Samuel; Croasdale, Rebecca; Trochanowska, Halina; Shao, Cuiying; Wang, Peng; Shi, Lei; Weinzierl, Tina; Rieder, Natascha; Bacac, Marina; Ries, Carola H; Kettenberger, Hubert; Schlothauer, Tilman; Friess, Thomas; Umana, Pablo; Klein, Christian

    2016-01-01

    The epidermal growth factor receptor (EGFR) and the insulin-like growth factor-1 receptor (IGF-1R) play critical roles in tumor growth, providing a strong rationale for the combined inhibition of IGF-1R and EGFR signaling in cancer therapy. We describe the design, affinity maturation, in vitro and in vivo characterization of the bispecific anti-IGF-1R/EGFR antibody XGFR*. XGFR* is based on the bispecific IgG antibody XGFR, which enabled heterodimerization of an IGF-1R binding scFab heavy chain with an EGFR-binding light and heavy chain by the "knobs-into-holes" technology. XGFR* is optimized for monovalent binding of human EGFR and IGF-1R with increased binding affinity for IGF-1R due to affinity maturation and highly improved protein stability to oxidative and thermal stress. It bears an afucosylated Fc-portion for optimal induction of antibody-dependent cell-mediated cytotoxicity (ADCC). Stable Chinese hamster ovary cell clones with production yields of 2-3 g/L were generated, allowing for large scale production of the bispecific antibody. XGFR* potently inhibits EGFR- and IGF-1R-dependent receptor phosphorylation, reduces tumor cell proliferation in cells with heterogeneous levels of IGF-1R and EGFR receptor expression and induces strong ADCC in vitro. A comparison of pancreatic and colorectal cancer lines demonstrated superior responsiveness to XGFR*-mediated signaling and tumor growth inhibition in pancreatic cancers that frequently show a high degree of IGF-1R/EGFR co-expression. XGFR* showed potent anti-tumoral efficacy in the orthotopic MiaPaCa-2 pancreatic xenograft model, resulting in nearly complete tumor growth inhibition with significant number of tumor remissions. In summary, the bispecific anti-IGF-1R/EGFR antibody XGFR* combines potent signaling and tumor growth inhibition with enhanced ADCC induction and represents a clinical development candidate for the treatment of pancreatic cancer. PMID:26984378

  8. Xanthate-Functional Temperature-Responsive Polymers: Effect on Lower Critical Solution Temperature Behavior and Affinity toward Sulfide Surfaces.

    Science.gov (United States)

    Ng, Wei Sung; Forbes, Elizaveta; Franks, George V; Connal, Luke A

    2016-08-01

    Xanthate-functional polymers represent an exciting opportunity to provide temperature-responsive materials with the ability to selectively attach to specific metals, while also modifying the lower critical solution temperature (LCST) behavior. To investigate this, random copolymers of poly(N-isopropylacrylamide) (PNIPAM) with xanthate incorporations ranging from 2 to 32% were prepared via free radical polymerization. Functionalization with 2% xanthate increased the LCST by 5 °C relative to the same polymer without xanthate. With increasing xanthate composition, the transition temperature increased and the transition range broadened until a critical composition of the hydrophilic xanthate groups (≥18%) where the transition disappeared completely. The adsorption of the polymers at room temperature onto chalcopyrite (CuFeS2) surfaces increased with xanthate composition, while adsorption onto quartz (SiO2) was negligible. These findings demonstrate the affinity of these functional smart polymers toward copper iron sulfide relative to quartz surfaces, presumably due to the interactions between xanthate and specific metal centers. PMID:27434760

  9. Mature high-affinity immune responses to (pro)insulin anticipate the autoimmune cascade that leads to type 1 diabetes

    OpenAIRE

    Achenbach, Peter; Koczwara, Kerstin; Knopff, Annette; Naserke, Heike; Ziegler, Anette-G.; Bonifacio, Ezio

    2004-01-01

    Children at risk for type 1 diabetes can develop early insulin autoantibodies (IAAs). Many, but not all, of these children subsequently develop multiple islet autoantibodies and diabetes. To determine whether disease progression is reflected by autoantibody maturity, IAA affinity was measured by competitive radiobinding assay in first and subsequent IAA-positive samples from children followed from birth in the BABYDIAB cohort. IAA affinity in first positive samples ranged from less than 106 l...

  10. Radiation responses of stem cells: targeted and non-targeted effects

    International Nuclear Information System (INIS)

    Stem cells are fundamental to the development of any tissue or organism via their ability to self-renew, which is aided by their unlimited proliferative capacity and their ability to produce fully differentiated offspring, often from multiple lineages. Stems cells are long lived and have the potential to accumulate mutations, including in response to radiation exposure. It is thought that stem cells have the potential to be induced into a cancer stem cell phenotype and that these may play an important role in resistance to radiotherapy. For radiation-induced carcinogenesis, the role of targeted and non-targeted effects is unclear with tissue or origin being important. Studies of genomic instability and bystander responses have shown consistent effects in haematopoietic models. Several models of radiation have predicted that stem cells play an important role in tumour initiation and that bystander responses could play a role in proliferation and self-renewal. (authors)

  11. Bystander responses in cells models; targets, dosimetry and mechanisms

    International Nuclear Information System (INIS)

    The use of microbeam approaches has been a major advance in probing the relevance of bystander responses in cell and tissue models. Our own studies at the Gray Cancer Institute have used both a charged particle microbeam, producing protons and helium ions and a soft X-ray microprobe, delivering focused carbon-K, aluminium-K and titanium-K soft X-rays. Using these techniques we have been able to build up a comprehensive picture of the underlying differences between bystander responses and direct effects in cell and tissue-like models. What is now clear is that bystander dose-response relationships, the underlying mechanisms of action and the targets involved are not the same as those observed for direct irradiation of DNA in the nucleus. Our recent studies have shown bystander responses induced in human or hamster cells even when radiation is deposited away from the nucleus in cytoplasmic targets either after charged particle or soft X-ray exposure. Importantly, the level of bystander effect, measured as cell killing was similar to that observed when the same amount of energy was deposited but targeted to the nucleus. In other studies, we have shown that underlying determination of the level of response is the energy deposited in a single cell rather than the number of cells hit. Also the overall response at low doses may be dominated by bystander signaling. These observations have significance for our understanding of radiation risk at low doses including those of environmental exposures and the applicability of the Linear Non Threshold model. The realization that cell to cell signaling is important for radiation response may also open up new therapeutic opportunities to either improve tumor cell kill or protect normal tissues if the pathways underpinning bystander signaling can be elucidated and controlled

  12. Curcumin binds in silico to anti-cancer drug target enzyme MMP-3 (human stromelysin-1) with affinity comparable to two known inhibitors of the enzyme.

    Science.gov (United States)

    Jerah, Ahmed; Hobani, Yahya; Kumar, B Vinod; Bidwai, Anil

    2015-01-01

    In silico interaction of curcumin with the enzyme MMP-3 (human stromelysin-1) was studied by molecular docking using AutoDock 4.2 as the docking software application. AutoDock 4.2 software serves as a valid and acceptable docking application to study the interactions of small compounds with proteins. Interactions of curcumin with MMP-3 were compared to those of two known inhibitors of the enzyme, PBSA and MPPT. The calculated free energy of binding (ΔG binding) shows that curcumin binds with affinity comparable to or better than the two known inhibitors. Binding interactions of curcumin with active site residues of the enzyme are also predicted. Curcumin appears to bind in an extendended conformation making extensive VDW contacts in the active site of the enzyme. Hydrogen bonding and pi-pi interactions with key active site residues is also observed. Thus, curcumin can be considered as a good lead compound in the development of new inhibitors of MMP-3 which is a potential target of anticancer drugs. The results of these studies can serve as a starting point for further computational and experimental studies. PMID:26420919

  13. Stimuli-responsive nanoparticles for targeting the tumor microenvironment.

    Science.gov (United States)

    Du, Jinzhi; Lane, Lucas A; Nie, Shuming

    2015-12-10

    One of the most challenging and clinically important goals in nanomedicine is to deliver imaging and therapeutic agents to solid tumors. Here we discuss the recent design and development of stimuli-responsive smart nanoparticles for targeting the common attributes of solid tumors such as their acidic and hypoxic microenvironments. This class of stimuli-responsive nanoparticles is inactive during blood circulation and under normal physiological conditions, but is activated by acidic pH, enzymatic up-regulation, or hypoxia once they extravasate into the tumor microenvironment. The nanoparticles are often designed to first "navigate" the body's vascular system, "dock" at the tumor sites, and then "activate" for action inside the tumor interstitial space. They combine the favorable biodistribution and pharmacokinetic properties of nanodelivery vehicles and the rapid diffusion and penetration properties of smaller drug cargos. By targeting the broad tumor habitats rather than tumor-specific receptors, this strategy has the potential to overcome the tumor heterogeneity problem and could be used to design diagnostic and therapeutic nanoparticles for a broad range of solid tumors. PMID:26341694

  14. Radiolabeled high affinity peptidomimetic antagonist selectively targets {alpha}{sub v}{beta}{sub 3} receptor-positive tumor in mice

    Energy Technology Data Exchange (ETDEWEB)

    Jang, Beom-Su [Department of Nuclear Medicine, Warren G. Magnuson Clinical Center, National Institutes of Health, Bethesda, MD 20892 (United States); Lim, Esther [Department of Radiology, Warren G. Magnuson Clinical Center, National Institutes of Health, Bethesda, MD 20892 (United States); Hee Park, Seung [Department of Nuclear Medicine, Warren G. Magnuson Clinical Center, National Institutes of Health, Bethesda, MD 20892 (United States); Shin, In Soo [Department of Nuclear Medicine, Warren G. Magnuson Clinical Center, National Institutes of Health, Bethesda, MD 20892 (United States); Danthi, S. Narasimhan [Department of Radiology, Warren G. Magnuson Clinical Center, National Institutes of Health, Bethesda, MD 20892 (United States); Hwang, In Sook [Department of Nuclear Medicine, Warren G. Magnuson Clinical Center, National Institutes of Health, Bethesda, MD 20892 (United States); Le, Nhat [Department of Nuclear Medicine, Warren G. Magnuson Clinical Center, National Institutes of Health, Bethesda, MD 20892 (United States); Yu, Sarah [Department of Nuclear Medicine, Warren G. Magnuson Clinical Center, National Institutes of Health, Bethesda, MD 20892 (United States); Xie Jianwu [Department of Radiology, Warren G. Magnuson Clinical Center, National Institutes of Health, Bethesda, MD 20892 (United States); Li, King C.P. [Department of Radiology, Warren G. Magnuson Clinical Center, National Institutes of Health, Bethesda, MD 20892 (United States); Carrasquillo, Jorge A. [Department of Nuclear Medicine, Warren G. Magnuson Clinical Center, National Institutes of Health, Bethesda, MD 20892 (United States); Paik, Chang H. [Department of Nuclear Medicine, Warren G. Magnuson Clinical Center, National Institutes of Health, Bethesda, MD 20892 (United States)]. E-mail: cpaik@mail.nih.gov

    2007-05-15

    Objectives: The aim of this research was to synthesize radiolabeled peptidomimetic integrin {alpha}{sub v}{beta}{sub 3} antagonists that selectively target integrin {alpha}{sub v}{beta}{sub 3} receptor and clear rapidly from the whole body. Methods: Integrin {alpha}{sub v}{beta}{sub 3} antagonists, 4-[2-(3,4,5,6-tetrahydropyrimidine-2-ylamino) ethyloxy]benzoyl-2-(S)-aminoethylsulfonyl-amino-{beta}-alanine (IA) and 4-[2-(3,4,5,6-tetrahydro-pyrimidin-2-ylamino)-ethyloxy]benzoyl-2-(S)-[N- (3-amino-neopenta-1-carbamyl)]-aminoethylsulfonylamino-{beta}-alanine hydrochloride (IAC), a hydrophobic carbamate derivative of IA, were conjugated with 2-p-isothiocyanatobenzyl-DOTA at the amino terminus and labeled with {sup 111}In. The {sup 111}In labeled IA and IAC were subjected to in vitro receptor binding, biodistribution and imaging studies using nude mice bearing the receptor-positive M21 human melanoma xenografts. Results: The {sup 111}In-labeled IA (40%) and -IAC (72%) specifically bound in vitro to {alpha}{sub v}{beta}{sub 3} (0.8 {mu}M) at a molar excess. This receptor binding was completely blocked by a molar excess of cold IA to {alpha}{sub v}{beta}{sub 3}. The higher receptor-binding affinity of the {sup 111}In-labeled IAC was reflected in higher tumor uptake and retention: 5.6{+-}1.4 and 4.5{+-}0.7 %ID/g vs. 3.8{+-}0.9 and 2.0{+-}0.3 %ID/g for the {sup 111}In-labeled IA at 0.33 and 2 h. The tumor uptakes were inhibited by the co-injection of 200 {mu}g of IA, indicating that the uptake was receptor mediated. These antagonists were excreted primarily via the renal system. The {sup 111}In activity retained in the whole body was quite comparable between the {sup 111}In-labeled IA (24% ID) and the {sup 111}In-labeled IAC (33% ID) at 2 h. The higher peak tumor uptake and longer retention resulted in higher tumor-to-background ratios for the {sup 111}In-labeled IAC at 2 h with 9.7, 2.3, 0.8, 1.9, 7.1, 2.2, 0.9, 3.7 and 9.9 for blood, liver, kidney, lung, heart, stomach

  15. Relacin, a novel antibacterial agent targeting the Stringent Response.

    Directory of Open Access Journals (Sweden)

    Ezequiel Wexselblatt

    2012-09-01

    Full Text Available Finding bacterial cellular targets for developing novel antibiotics has become a major challenge in fighting resistant pathogenic bacteria. We present a novel compound, Relacin, designed to inhibit (pppGpp production by the ubiquitous bacterial enzyme RelA that triggers the Stringent Response. Relacin inhibits RelA in vitro and reduces (pppGpp production in vivo. Moreover, Relacin affects entry into stationary phase in Gram positive bacteria, leading to a dramatic reduction in cell viability. When Relacin is added to sporulating Bacillus subtilis cells, it strongly perturbs spore formation regardless of the time of addition. Spore formation is also impeded in the pathogenic bacterium Bacillus anthracis that causes the acute anthrax disease. Finally, the formation of multicellular biofilms is markedly disrupted by Relacin. Thus, we establish that Relacin, a novel ppGpp analogue, interferes with bacterial long term survival strategies, placing it as an attractive new antibacterial agent.

  16. A versatile fluorescent biosensor based on target-responsive graphene oxide hydrogel for antibiotic detection.

    Science.gov (United States)

    Tan, Bing; Zhao, Huimin; Du, Lei; Gan, Xiaorong; Quan, Xie

    2016-09-15

    A fluorescent sensing platform based on graphene oxide (GO) hydrogel was developed through a fast and facile gelation, immersion and fluorescence determination process, in which the adenosine and aptamer worked as the co-crosslinkers to connect the GO sheets and then form the three-dimensional (3D) macrostructures. The as-prepared hydrogel showed high mechanical strength and thermal stability. The optimal hydrogel had a linear response for oxytetracycline (OTC) of 25-1000μg/L and a limit of quantitation (LOQ) of 25μg/L. Moreover, together with the high affinity of the aptamer for its target, this assay exhibited excellent sensitivity and selectivity. According to its design principle, the as-designed hydrogel was also tested to possess the generic detection function for other molecules by simply replacing its recognition element, which is expected to lay a foundation to realize the assembly of functionalized hierarchical graphene-based materials for practical applications in analytical field. PMID:27132000

  17. Stimuli-responsive PEGylated prodrugs for targeted doxorubicin delivery

    Energy Technology Data Exchange (ETDEWEB)

    Xu, Minghui; Qian, Junmin, E-mail: jmqian@mail.xjtu.edu.cn; Liu, Xuefeng; Liu, Ting; Wang, Hongjie

    2015-05-01

    In recent years, stimuli-sensitive prodrugs have been extensively studied for the rapid “burst” release of antitumor drugs to enhance chemotherapeutic efficiency. In this study, a novel stimuli-sensitive prodrug containing galactosamine as a targeting moiety, poly(ethylene glycol)–doxorubicin (PEG–DOX) conjugate, was developed for targeting HepG2 human liver cancer cells. To obtain the PEG–DOX conjugate, both galactosamine-decorated poly(ethylene glycol) aldehyde (Gal-PEG-CHO) and methoxy poly(ethylene glycol) aldehyde (mPEG-CHO) were firstly synthesized and functionalized with dithiodipropionate dihydrazide (TPH) through direct reductive amination via Schiff's base formation, and then DOX molecules were chemically conjugated to the hydrazide end groups of TPH-functionalized Gal-/m-PEG chains via pH-sensitive hydrazone linkages. The chemical structures of TPH-functionalized PEG and PEG–DOX prodrug were confirmed by {sup 1}H NMR analysis. The PEG–DOX conjugate could self-assemble into spherical nanomicelles with a mean diameter of 140 nm, as indicated by transmission electron microscopy and dynamic light scattering. The drug loading content and loading efficiency in the prodrug nanomicelles were as high as 20 wt.% and 75 wt.%, respectively. In vitro drug release studies showed that DOX was released rapidly from the prodrug nanomicelles at the intracellular levels of pH and reducing agent. Cellular uptake and MTT experiments demonstrated that the galactosamine-decorated prodrug nanomicelles were more efficiently internalized into HepG2 cells via a receptor-mediated endocytosis process and exhibited a higher toxicity, compared with pristine prodrug nanomicelles. These results suggest that the novel Gal-PEG–DOX prodrug nanomicelles have tremendous potential for targeted liver cancer therapy. - Highlights: • A novel stimuli-responsive PEGylated prodrugs is synthesized. • PEGylated prodrugs can self-assemble into spherical nanoparticles (140 nm

  18. Monocarboxylate transporters as targets and mediators in cancer therapy response.

    Science.gov (United States)

    Baltazar, F; Pinheiro, C; Morais-Santos, F; Azevedo-Silva, J; Queirós, O; Preto, A; Casal, M

    2014-12-01

    Monocarboxylate transporters (MCTs) belong to a family of transporters, encoded by the SLC16 gene family, which is presently composed by 14 members, but only MCT1 to 4 have been biochemically characterized. They have important functions in healthy tissues, being involved in the transmembrane transport of lactic acid and other monocarboxylic acids in human cells. One of the recently recognized hallmarks of cancer is altered metabolism, with high rates of glucose consumption and consequent lactate production. To maintain this metabolic phenotype, cancer cells upregulate a series of plasma membrane proteins, including MCTs. MCT1 and MCT4, in particular, play a dual role in the maintenance of the metabolic phenotype of tumour cells. On one hand, they facilitate the efflux of lactate and, on the other hand, they contribute to the preservation of the intracellular pH, by co-transporting a proton. Thus, MCTs are attractive targets in cancer therapy, especially in cancers with a hyper-glycolytic and acid-resistant phenotype. Recent evidence demonstrates that MCTs are involved in cancer cell uptake of chemotherapeutic agents, including 3-bromopyruvate. In this way MCTs can act as "Trojan horses", as their elevated expression in cancer cells can mediate the entry of this chemotherapeutic agent into the cells and selectively kill cancer cells. As a result, MCTs will be mediators of chemotherapeutic response, and their expression can be used as a molecular marker to predict response to chemotherapy. PMID:24921258

  19. Protein Complex Purification by Affinity Capture.

    Science.gov (United States)

    LaCava, John; Fernandez-Martinez, Javier; Hakhverdyan, Zhanna; Rout, Michael P

    2016-01-01

    Affinity capture has become a powerful technique for consistently purifying endogenous protein complexes, facilitating biochemical and biophysical assays on otherwise inaccessible biological assemblies, and enabling broader interactomic exploration. For this procedure, cells are broken and their contents separated and extracted into a solvent, permitting access to target macromolecular complexes thus released in solution. The complexes are specifically enriched from the extract onto a solid medium coupled with an affinity reagent-usually an antibody-that recognizes the target either directly or through an appended affinity tag, allowing subsequent characterization of the complex. Here, we discuss approaches and considerations for purifying endogenous yeast protein complexes by affinity capture. PMID:27371601

  20. Real-time detection of implant-associated neutrophil responses using a formyl peptide receptor-targeting NIR nanoprobe

    Directory of Open Access Journals (Sweden)

    Zhou J

    2012-05-01

    Full Text Available Jun Zhou*, Yi-Ting Tsai*, Hong Weng, Ewin N Tang, Ashwin Nair, Digant P Davé, Liping Tang, Department of Bioengineering, University of Texas at Arlington, Arlington, TX *Both authors contributed equally to this workAbstract: Neutrophils play an important role in implant-mediated inflammation and infection. Unfortunately, current methods which monitor neutrophil activity, including enzyme measurements and histological evaluation, require many animals and cannot be used to accurately depict the dynamic cellular responses. To understand the neutrophil interactions around implant-mediated inflammation and infection it is critical to develop methods which can monitor in vivo cellular activity in real time. In this study, formyl peptide receptor (FPR-targeting near-infrared nanoprobes were fabricated. This was accomplished by conjugating near-infrared dye with specific peptides having a high affinity to the FPRs present on activated neutrophils. The ability of FPR-targeting nanoprobes to detect and quantify activated neutrophils was assessed both in vitro and in vivo. As expected, FPR-targeting nanoprobes preferentially accumulated on activated neutrophils in vitro. Following transplantation, FPR-targeting nanoprobes preferentially accumulated at the biomaterial implantation site. Equally important, a strong relationship was observed between the extent of fluorescence intensity in vivo and the number of recruited neutrophils at the implantation site. Furthermore, FPR-targeting nanoprobes may be used to detect and quantify the number of neutrophils responding to a catheter-associated infection. The results show that FPR-targeting nanoprobes may serve as a powerful tool to monitor and measure the extent of neutrophil responses to biomaterial implants in vivo.Keywords: in vivo imaging, nanoprobe, neutrophils, inflammation, biocompatibility

  1. Magnetically responsive microparticles for targeted drug and radionuclide delivery.

    Energy Technology Data Exchange (ETDEWEB)

    Kaminski, M. D.; Ghebremeskel, A. N.; Nunez, L.; Kasza, K. E.; Chang, F.; Chien, T.-H.; Fisher, P. F.; Eastman, J. A.; Rosengart, A. J.; McDonald, L.; Xie, Y.; Johns, L.; Pytel, P.; Hafeli, U. O.

    2004-02-16

    We are currently investigating the use of magnetic particles--polymeric-based spheres containing dispersed magnetic nanocrystalline phases--for the precise delivery of drugs via the human vasculature. According to this review, meticulously prepared magnetic drug targeting holds promise as a safe and effective method of delivering drugs to specific organ, tissue or cellular targets. We have critically examined the wide range of approaches in the design and implementation of magnetic-particle-based drug delivery systems to date, including magnetic particle preparation, drug encapsulation, biostability, biocompatibility, toxicity, magnetic field designs, and clinical trials. However, we strongly believe that there are several limitations with past developments that need to be addressed to enable significant strides in the field. First, particle size has to be carefully chosen. Micrometer-sized magnetic particles are better attracted over a distance than nanometer sized magnetic particles by a constant magnetic field gradient, and particle sizes up to 1 {micro}m show a much better accumulation with no apparent side effects in small animal models, since the smallest blood vessels have an inner diameter of 5-7 {micro}m. Nanometer-sized particles <70 nm will accumulate in organ fenestrations despite an effective surface stabilizer. To be suitable for future human applications, our experimental approach synthesizes the magnetic drug carrier according to specific predefined outcome metrics: monodisperse population in a size range of 100 nm to 1.0 {micro}m, non-toxic, with appropriate magnetic properties, and demonstrating successful in vitro and in vivo tests. Another important variable offering possible improvement is surface polarity, which is expected to prolong particle half-life in circulation and modify biodistribution and stability of drugs in the body. The molecules in the blood that are responsible for enhancing the uptake of particles by the reticuloendothelial

  2. Class II-targeted antigen is superior to CD40-targeted antigen at stimulating humoral responses in vivo.

    Science.gov (United States)

    Frleta, D; Demian, D; Wade, W F

    2001-02-01

    We examined the efficacy of using monoclonal antibodies to target antigen (avidin) to different surface molecules expressed on antigen presenting cells (APC). In particular, we targeted CD40 to test whether the "adjuvant" properties of CD40 signaling combined with targeted antigen would result in enhanced serologic responses. We targeted avidin to class II as a positive control and to CD11c as a negative control. These surface proteins represent an ensemble of surface molecules that signal upon ligation and that are expressed on professional APC, in particular dendritic cells (DC). We observed that targeting class II molecules on APC was superior to targeting CD40, or CD11c. However, CD40 and CD11c could function as targets for antigen bound monoclonal antibodies under certain conditions. Interestingly, inclusion of anti-CD40 mAb with the targeting anti-class II-targeted antigens negatively affects humoral response, suggesting that CD40 signaling under certain conditions may suppress processing and/or presentation of targeted antigen. PMID:11360928

  3. Methods for Improving Aptamer Binding Affinity

    OpenAIRE

    Hijiri Hasegawa; Nasa Savory; Koichi Abe; Kazunori Ikebukuro

    2016-01-01

    Aptamers are single stranded oligonucleotides that bind a wide range of biological targets. Although aptamers can be isolated from pools of random sequence oligonucleotides using affinity-based selection, aptamers with high affinities are not always obtained. Therefore, further refinement of aptamers is required to achieve desired binding affinities. The optimization of primary sequences and stabilization of aptamer conformations are the main approaches to refining the binding properties of a...

  4. HAMS: High-Affinity Mass Spectrometry Screening. A High-Throughput Screening Method for Identifying the Tightest-Binding Lead Compounds for Target Proteins with No False Positive Identifications

    Science.gov (United States)

    Imaduwage, Kasun P.; Go, Eden P.; Zhu, Zhikai; Desaire, Heather

    2016-09-01

    A major challenge in drug discovery is the identification of high affinity lead compounds that bind a particular target protein; these leads are typically identified by high throughput screens. Mass spectrometry has become a detection method of choice in drug screening assays because the target and the ligand need not be modified. Label-free assays are advantageous because they can be developed more rapidly than assays requiring labels, and they eliminate the risk of the label interfering with the binding event. However, in commonly used MS-based screening methods, detection of false positives is a major challenge. Here, we describe a detection strategy designed to eliminate false positives. In this approach, the protein and the ligands are incubated together, and the non-binders are separated for detection. Hits (protein binders) are not detectable by MS after incubation with the protein, but readily identifiable by MS when the target protein is not present in the incubation media. The assay was demonstrated using three different proteins and hundreds of non-inhibitors; no false positive hits were identified in any experiment. The assay can be tuned to select for ligands of a particular binding affinity by varying the quantity of protein used and the immobilization method. As examples, the method selectively detected inhibitors that have Ki values of 0.2 μM, 50 pM, and 700 pM. These findings demonstrate that the approach described here compares favorably with traditional MS-based screening methods.

  5. Dynamic response of the target container under pulsed heating

    Energy Technology Data Exchange (ETDEWEB)

    Liping Ni [Paul Scherrer Inst. (PSI), Villigen (Switzerland)

    1997-09-01

    The structural mechanics of a liquid target container for pulsed spallation sources have been simulated using both a commercial code and a PSI-developed program. Results from the transient thermal-structural analysis showed that, due to inertia effects, the dynamic stress in the target container is contributed mainly from direct heating in the initial time stage, and later from the pressure wave in the target liquid once it reaches the wall. (author) figs., tab., refs.

  6. Expression of Brassica napus L. γ-Glutamylcysteine Synthetase and Low-and High-Affinity Sulfate Transporters in Response to Excess Cadmium

    Institute of Scientific and Technical Information of China (English)

    Xin SUN; Xue-Mei SUN; Zhi-Min YANG; Shao-Qiong LI; Jin WANG; Song-Hua WANG

    2005-01-01

    In both the roots and leaves ofBrassica napus L. cv. Youyan No. 8 under treatment with 30 μmol/L Cd, massive production of non-protein thiols (NPT; mainly containing glutathione (GSH) and phytochelatins (PCs)) was induced, together with an increase in γ-glutamylcysteine synthetase (γ-ECS)mRNA transcripts. Because γ-ECS is the key enzyme catalyzing the first step in GSH biosynthesis, which, in turn, is converted to PCs, the Cd-induced increase in γ-ECS expression may be responsible for the observed increase in the production of NPT. Using a quantitative reverse transcription polymerase chain reaction (RT-PCR) approach, the expression of genes encoding a putative low-affinity sulfate transporter (LAST) and a putative high-affinity sulfate transporter (HAST) was determined at the transcriptional level. The RT-PCR analysis of relative transcript amounts indicates that the LAST gene in B. napus leaves showed a constitutive expression, which was hardly affected by Cd treatment. However, treatment with 30 μmol/L Cd for 2 or 3 d induced a marked increase in the expression of LAST in roots. Transcriptional expression of the HAST gene occurred in roots, but not in leaves. The expression of HAST only in the roots suggests that it has a specific function in sulfate uptake from soil and that the putative LAST may be responsible for the transport of sulfate from the roots to the shoots, as well as for the uptake of sulfate from soil. These results indicate that changes in transcriptional expression for sulfate transporters were required for the increased demand for sulfate during Cd stress.

  7. Specificity of Bacillus thuringiensis endotoxins is correlated with the presence of high-affinity binding sites in the brush border membrane of target insect midguts

    International Nuclear Information System (INIS)

    Binding studies were performed with two 125I-labeled Bacillus thuringiensis δ-endotoxins on brush border membrane vesicles prepared from the larval midgut of the tobacco hornworm Manduca sexta or the cabbage butterfly Pieris brassicae. One δ-endotoxin, Bt2-protoxin, is a 130-kDa recombinant crystalline protein from B. thuringiensis subsp. berliner. It kills larvae of both insect species. The active Bt2-toxin is a 60-kDa proteolytic fragment of the Bt2-protoxin. It binds saturably and with high affinity to brush border membrane vesicles from the midgut of both species. The other δ-endotoxin, Bt4412-protoxin, is a 136-kDa crystalline protein from B. thuringiensis subsp. thuringiensis, which is highly toxic for P. brassicae, but not for M. sexta larvae. Bt4412-toxin, obtained after proteolytic activation of Bt4412-protoxin, shows high-affinity saturable binding to P. brassicae vesicles but not to M. sexta vesicles. The correlation between toxicity and specific binding is further strengthened by competition studies. Other B. thuringiensis δ-endotoxins active against M. sexta compete for binding of 125I-labeled Bt2-toxin to M. sexta vesicles, whereas toxins active against dipteran or coleopteran larvae do not compete. Bt2-toxin and Bt4412-toxin bind to different sites on P. brassicae vesicles

  8. Specificity of Bacillus thuringiensis endotoxins is correlated with the presence of high-affinity binding sites in the brush border membrane of target insect midguts

    Energy Technology Data Exchange (ETDEWEB)

    Hofmann, C.; Vanderbruggen, H.; Hoefte, H.; Van Rie, J.; Jansens, S.; Van Mellaert, H. (J. Plateaustraat, Gent (Belgium))

    1988-11-01

    Binding studies were performed with two {sup 125}I-labeled Bacillus thuringiensis {delta}-endotoxins on brush border membrane vesicles prepared from the larval midgut of the tobacco hornworm Manduca sexta or the cabbage butterfly Pieris brassicae. One {delta}-endotoxin, Bt2-protoxin, is a 130-kDa recombinant crystalline protein from B. thuringiensis subsp. berliner. It kills larvae of both insect species. The active Bt2-toxin is a 60-kDa proteolytic fragment of the Bt2-protoxin. It binds saturably and with high affinity to brush border membrane vesicles from the midgut of both species. The other {delta}-endotoxin, Bt4412-protoxin, is a 136-kDa crystalline protein from B. thuringiensis subsp. thuringiensis, which is highly toxic for P. brassicae, but not for M. sexta larvae. Bt4412-toxin, obtained after proteolytic activation of Bt4412-protoxin, shows high-affinity saturable binding to P. brassicae vesicles but not to M. sexta vesicles. The correlation between toxicity and specific binding is further strengthened by competition studies. Other B. thuringiensis {delta}-endotoxins active against M. sexta compete for binding of {sup 125}I-labeled Bt2-toxin to M. sexta vesicles, whereas toxins active against dipteran or coleopteran larvae do not compete. Bt2-toxin and Bt4412-toxin bind to different sites on P. brassicae vesicles.

  9. Structural dynamic response of target container against proton beam

    Energy Technology Data Exchange (ETDEWEB)

    Kikuchi, Kenji; Ishikura, Syuichi; Futakawa, Masatoshi; Hino, Ryutaro [Japan Atomic Energy Research Inst., Tokai, Ibaraki (Japan). Tokai Research Establishment

    1997-11-01

    Stress waves were analyzed for a target container of neutron science research project using a high-intensity proton accelerator that generates high energy and high current proton beam. In the mercury target, the pulsed proton beam generates intense power density in the course of spallation reaction and causes pressure wave in the mercury and stress wave in the target container due to a sudden temperature change. Structural integrity of the target container depends on the power intensity at a maximum energy deposit. A broad proton profile is favorable to the structural assessment of the container rather than narrow one. Stress wave have propagated in the target container at a speed of sound. It only takes 0.1 ms for the size of 40 cm length stainless steel container. Further assessment is necessary to optimize a geometry of the container and establish a method to evaluate a life time. (author)

  10. Examination of Studies Targeting Social Skills with Pivotal Response Treatment

    Science.gov (United States)

    Bozkus Genc, Gulden; Vuran, Sezgin

    2013-01-01

    In early education, especially in effective teaching to children with autism spectrum disorders, the teaching methods which are applicable in natural settings like pivotal response treatment (PRT) are commonly used. It is one of the naturalistic intervention models aiming to facilitate the stimulant-response generalization, decrease the dependency…

  11. Polarizing T and B cell responses by APC-targeted subunit vaccines.

    Directory of Open Access Journals (Sweden)

    Gunnveig eGrødeland

    2015-07-01

    Full Text Available Current influenza vaccines mostly aim at the induction of specific neutralizing antibodies. While antibodies are important for protection against a particular virus strain, T cells can recognize epitopes that will offer broader protection against influenza. We have previously developed a DNA vaccine format by which protein antigens can be targeted specifically to receptors on antigen presenting cells (APCs. The DNA-encoded vaccine proteins are homodimers, each chain consisting of a targeting unit, a dimerization unit, and an antigen. The strategy of targeting antigen to APCs greatly enhances immune responses as compared to non-targeted controls. Furthermore, targeting of antigen to different receptors on APCs can polarize the immune response to different arms of immunity. Here, we discuss how targeting of hemagglutinin (HA to MHC class II molecules increases Th2 and IgG1 antibody responses, whereas targeting to chemokine receptors XCR1 or CCR1/3/5 increases Th1 and IgG2a responses, in addition to CD8+ T cell responses. We also discuss these results in relation to work published by others on APC-targeting. Differential targeting of APC surface molecules may allow the induction of tailor-made phenotypes of adaptive immune responses that are optimal for protection against various infectious agents, including influenza virus.

  12. Nonintentional behavioural responses to psi : hidden targets and hidden observers

    OpenAIRE

    Anderson, Mary-Jane Charlotte

    2012-01-01

    Psi is the phenomenon of apparently responding to or receiving information by means other than the recognised senses. Psi information may influence human behaviour, without the individual intending this or even being aware of it. This thesis seeks to investigate nonintentional behavioural responses to psi. We present five empirical studies that investigated nonintentional behavioural responses to psi information. In each study, the psi information was hidden from participants, ...

  13. Stable Extended Human Immunodeficiency Virus Type 1 gp41 Coiled Coil as an Effective Target in an Assay for High-Affinity Fusion Inhibitors▿

    OpenAIRE

    Cai, Lifeng; Balogh, Edina; Gochin, Miriam

    2009-01-01

    The human immunodeficiency virus type 1 (HIV-1) gp41 coiled-coil domain is an important target for fusion inhibitors, including the peptide T20, which has been approved as a drug against HIV-1. Research into nonpeptide fusion inhibitors has focused primarily on a hydrophobic pocket located within the coiled coil and has so far yielded compounds with relatively weak fusion inhibitory activity. Here, we describe metal ion-assisted stabilization of an extended 39-residue construct of gp41, which...

  14. Is It Reliable to Use Common Molecular Docking Methods for Comparing the Binding Affinities of Enantiomer Pairs for Their Protein Target?

    Science.gov (United States)

    Ramírez, David; Caballero, Julio

    2016-01-01

    Molecular docking is a computational chemistry method which has become essential for the rational drug design process. In this context, it has had great impact as a successful tool for the study of ligand-receptor interaction modes, and for the exploration of large chemical datasets through virtual screening experiments. Despite their unquestionable merits, docking methods are not reliable for predicting binding energies due to the simple scoring functions they use. However, comparisons between two or three complexes using the predicted binding energies as a criterion are commonly found in the literature. In the present work we tested how wise is it to trust the docking energies when two complexes between a target protein and enantiomer pairs are compared. For this purpose, a ligand library composed by 141 enantiomeric pairs was used, including compounds with biological activities reported against seven protein targets. Docking results using the software Glide (considering extra precision (XP), standard precision (SP), and high-throughput virtual screening (HTVS) modes) and AutoDock Vina were compared with the reported biological activities using a classification scheme. Our test failed for all modes and targets, demonstrating that an accurate prediction when binding energies of enantiomers are compared using docking may be due to chance. We also compared pairs of compounds with different molecular weights and found the same results. PMID:27104528

  15. Know your epidemic, know your response: targeting HIV in Asia

    DEFF Research Database (Denmark)

    Lazarus, Jeff; Curth, Nadja; Bridge, Jamie;

    2010-01-01

    This article provides an overview of the HIV epidemic in Asia, the context within which the epidemic is evolving, and the key actions to address the challenges faced by countries and risk groups. HIV epidemics across Asia are predominantly concentrated among most-at-risk populations. Although...... prevention and treatment services. In order to reach the Millennium Development Goal of halting and reversing the spread of HIV by 2015 and to achieve universal access to HIV treatment, these barriers must be overcome across Asia. High-impact programs must be targeted at those in need, with continuous...

  16. Detrimental effects of albuterol on airway responsiveness requires airway inflammation and is independent of β-receptor affinity in murine models of asthma

    Directory of Open Access Journals (Sweden)

    Aimi Steven

    2011-03-01

    Full Text Available Abstract Background Inhaled short acting β2-agonists (SABA, e.g. albuterol, are used for quick reversal of bronchoconstriction in asthmatics. While SABA are not recommended for maintenance therapy, it is not uncommon to find patients who frequently use SABA over a long period of time and there is a suspicion that long term exposure to SABA could be detrimental to lung function. To test this hypothesis we studied the effect of long-term inhaled albuterol stereoisomers on immediate allergic response (IAR and airway hyperresponsiveness (AHR in mouse models of asthma. Methods Balb/C mice were sensitized and challenged with ovalbumin (OVA and then we studied the IAR to inhaled allergen and the AHR to inhaled methacholine. The mice were pretreated with nebulizations of either racemic (RS-albuterol or the single isomers (S- and (R-albuterol twice daily over 7 days prior to harvest. Results We found that all forms of albuterol produced a significant increase of IAR measured as respiratory elastance. Similarly, we found that AHR was elevated by albuterol. At the same time a mouse strain that is intrinsically hyperresponsive (A/J mouse was not affected by the albuterol isomers nor was AHR induced by epithelial disruption with Poly-L-lysine affected by albuterol. Conclusions We conclude that long term inhalation treatment with either isomer of albuterol is capable of precipitating IAR and AHR in allergically inflamed airways but not in intrinsically hyperresponsive mice or immunologically naïve mice. Because (S-albuterol, which lacks affinity for the β2-receptor, did not differ from (R-albuterol, we speculate that isomer-independent properties of the albuterol molecule, other than β2-agonism, are responsible for the effect on AHR.

  17. 具有葡萄糖转运蛋白亲和力的脑靶向文拉法辛前药的合成%Synthesis of brain-targeting prodrug of Venlafaxine with affinity to GLUT1

    Institute of Scientific and Technical Information of China (English)

    张勇; 吴雪莹; 任云; 海俐

    2013-01-01

    OBJECTIVE To design and synthesize a brain-targeting prodrug of Venlafaxine with affinity to the glucose transporter 1 (GLUT1).METHODS Venlafaxine,as starting material,was conjugated with 1,2,3,4-tetra-O-trimethysilyl-D-glucopyranose by succinic acid,and then deprotected to get the target compound Ⅰ.RESULTS and CONCLUSION The target compound was confirmed by 1HNMR,IR and MS.%目的 为了提高抗抑郁药文拉法辛的脑靶向性,以血脑屏障上的葡萄糖转运蛋白1为靶标,设计并合成了脑靶向性的文拉法辛前药Ⅰ.方法 以文拉法辛为起始原料,通过丁二酸为桥链与三甲硅基保护的葡萄糖偶联,脱去保护基,得到Ⅰ.结果和结论 目标化合物Ⅰ经1HNMR、IR和MS确证结构.

  18. Dual responsive PNIPAM–chitosan targeted magnetic nanopolymers for targeted drug delivery

    Energy Technology Data Exchange (ETDEWEB)

    Yadavalli, Tejabhiram, E-mail: tejabhiram@gmail.com [Nanotechnology Research Centre, SRM University, Chennai 603203 (India); Ramasamy, Shivaraman [Nanotechnology Research Centre, SRM University, Chennai 603203 (India); School of Physics, The University of Western Australia, 35 Stirling Hwy, Crawley, WA 6009 (Australia); Chandrasekaran, Gopalakrishnan; Michael, Isaac; Therese, Helen Annal [Nanotechnology Research Centre, SRM University, Chennai 603203 (India); Chennakesavulu, Ramasamy [Department of Pharmacy practice, SRM College of Pharmacy, Chennai 603203 (India)

    2015-04-15

    A dual stimuli sensitive magnetic hyperthermia based drug delivery system has been developed for targeted cancer treatment. Thermosensitive amine terminated poly-N-isopropylacrylamide complexed with pH sensitive chitosan nanoparticles was prepared as the drug carrier. Folic acid and fluorescein were tagged to the nanopolymer complex via N-hydroxysuccinimide and ethyl-3-(3-dimethylaminopropyl)carbodiimide reaction to form a fluorescent and cancer targeting magnetic carrier system. The formation of the polymer complex was confirmed using infrared spectroscopy. Gadolinium doped nickel ferrite nanoparticles prepared by a hydrothermal method were encapsulated in the polymer complex to form a magnetic drug carrier system. The proton relaxation studies on the magnetic carrier system revealed a 200% increase in the T1 proton relaxation rate. These magnetic carriers were loaded with curcumin using solvent evaporation method with a drug loading efficiency of 86%. Drug loaded nanoparticles were tested for their targeting and anticancer properties on four cancer cell lines with the help of MTT assay. The results indicated apoptosis of cancer cell lines within 3 h of incubation. - Highlights: • The use of gadolinium doped nickel ferrite with the suggested doping level. • The use of PNIPMA–chitosan polymer with folic acid and fluorescein as a drug carrier complex. • Magnetic hyperthermia studies of gadolinium doped nickel ferrites are being reported for the first time. • Proton relaxivity studies which indicate the MRI contrasting properties on the reported system are new. • Use of curcumin, a hydrophobic Indian spice as a cancer killing agent inside the reported magnetic polymer complex.

  19. Dual responsive PNIPAM–chitosan targeted magnetic nanopolymers for targeted drug delivery

    International Nuclear Information System (INIS)

    A dual stimuli sensitive magnetic hyperthermia based drug delivery system has been developed for targeted cancer treatment. Thermosensitive amine terminated poly-N-isopropylacrylamide complexed with pH sensitive chitosan nanoparticles was prepared as the drug carrier. Folic acid and fluorescein were tagged to the nanopolymer complex via N-hydroxysuccinimide and ethyl-3-(3-dimethylaminopropyl)carbodiimide reaction to form a fluorescent and cancer targeting magnetic carrier system. The formation of the polymer complex was confirmed using infrared spectroscopy. Gadolinium doped nickel ferrite nanoparticles prepared by a hydrothermal method were encapsulated in the polymer complex to form a magnetic drug carrier system. The proton relaxation studies on the magnetic carrier system revealed a 200% increase in the T1 proton relaxation rate. These magnetic carriers were loaded with curcumin using solvent evaporation method with a drug loading efficiency of 86%. Drug loaded nanoparticles were tested for their targeting and anticancer properties on four cancer cell lines with the help of MTT assay. The results indicated apoptosis of cancer cell lines within 3 h of incubation. - Highlights: • The use of gadolinium doped nickel ferrite with the suggested doping level. • The use of PNIPMA–chitosan polymer with folic acid and fluorescein as a drug carrier complex. • Magnetic hyperthermia studies of gadolinium doped nickel ferrites are being reported for the first time. • Proton relaxivity studies which indicate the MRI contrasting properties on the reported system are new. • Use of curcumin, a hydrophobic Indian spice as a cancer killing agent inside the reported magnetic polymer complex

  20. Bisphosphonates target B cells to enhance humoral immune responses

    OpenAIRE

    Tonti, Elena; Jiménez de Oya, Nereida; Galliverti, Gabriele; Moseman, E. Ashley; Di Lucia, Pietro; Amabile, Angelo; Sammicheli, Stefano; De Giovanni, Marco; Sironi, Laura; Chevrier, Nicolas; Sitia, Giovanni; Gennari, Luigi; Guidotti, Luca G.; von Andrian, Ulrich H.; Iannacone, Matteo

    2013-01-01

    Bisphosphonates are a class of drugs that are widely used to inhibit loss of bone mass in patients. We show here that the administration of clinically relevant doses of bisphosphonates in mice increases antibody responses to live and inactive viruses, proteins, haptens and existing commercial vaccine formulations. Bisphosphonates exert this adjuvant-like activity in the absence of CD4+ and γδ T cells, neutrophils or dendritic cells and their effect does not rely on local macrophage depletion ...

  1. Innate inflammatory responses in stroke: mechanisms and potential therapeutic targets

    OpenAIRE

    Kim, Jong Youl; Kawabori, Masahito; Yenari, Midori A.

    2014-01-01

    Stroke is a frequent cause of long-term disability and death worldwide. Ischemic stroke is more commonly encountered compared to hemorrhagic stroke, and leads to tissue death by ischemia due to occlusion of a cerebral artery. Inflammation is known to result as a result of ischemic injury, long thought to be involved in initiating the recovery and repair process. However, work over the past few decades indicates that aspects of this inflammatory response may in fact be detrimental to stroke ou...

  2. Affine Grassmann codes

    DEFF Research Database (Denmark)

    Høholdt, Tom; Beelen, Peter; Ghorpade, Sudhir Ramakant

    2010-01-01

    We consider a new class of linear codes, called affine Grassmann codes. These can be viewed as a variant of generalized Reed-Muller codes and are closely related to Grassmann codes.We determine the length, dimension, and the minimum distance of any affine Grassmann code. Moreover, we show that...... affine Grassmann codes have a large automorphism group and determine the number of minimum weight codewords....

  3. Synthesis and biological evaluation of copper-64 radiolabeled [DUPA-6-Ahx-(NODAGA)-5-Ava-BBN(7-14)NH2], a novel bivalent targeting vector having affinity for two distinct biomarkers (GRPr/PSMA) of prostate cancer

    International Nuclear Information System (INIS)

    Gastrin-releasing peptide receptors (GRPr) and prostate-specific membrane antigen (PSMA) are two identifying biomarkers expressed in very high numbers on prostate cancer cells and could serve as a useful tool for molecular targeting and diagnosis of disease via positron-emission tomography (PET). The aim of this study was to produce the multipurpose, bivalent [DUPA-6-Ahx-(64Cu-NODAGA)-5-Ava-BBN(7-14)NH2] radioligand for prostate cancer imaging, where DUPA = (2-[3-(1,3-dicarboxypropyl)-ureido]pentanedioic acid), a small-molecule, PSMA-targeting probe, 6Ahx = 6-aminohexanoic acid, 5-Ava = 5-aminovaleric acid, NODAGA = [2-(4,7-biscarboxymethyl)-1,4,7-(triazonan-1-yl)pentanedioic acid] (a derivative of NOTA (1,4,7-triazacyclononane-1,4,7-triacetic acid)), and BBN(7-14)NH2 = bombesin, a GRPr-specific peptide targeting probe. Methods: The PSMA/GRPr dual targeting ligand precursor [DUPA-6-Ahx-K-5-Ava-BBN(7-14)NH2], was synthesized by solid-phase and manual peptide synthesis, after which NODAGA was added via manual conjugation to the ε-amine of lysine (K). The new bivalent GRPr/PSMA targeting vector was purified by reversed-phase high performance liquid chromatography (RP-HPLC), characterized by electrospray-ionization mass spectrometry (ESI-MS), and metallated with 64CuCl2 and natCuCl2. The receptor binding affinity was evaluated in human, prostate, PC-3 (GRPr-positive) and LNCaP (PSMA-positive) cells and the tumor-targeting efficacy determined in severe combined immunodeficient (SCID) and athymic nude mice bearing PC-3 and LNCaP tumors. Whole-body maximum intensity microPET/CT images of PC-3/LNCaP tumor-bearing mice were obtained 18 h post-injection (p.i.). Results: Competitive binding assays in PC-3 and LNCaP cells indicated high receptor binding affinity for the [DUPA-6-Ahx-(natCu-NODAGA)-5-Ava-BBN(7-14)NH2] conjugate. MicroPET scintigraphy in PC-3/LNCaP tumor-bearing mice indicated that xenografted tumors were visible at 18 h p.i. with collateral, background

  4. Decreasing affinity of CAR-T cells targeting HER2 to increase therapeutic out-come against tumors%降低亲和力提高HER2-CAR-T细胞治疗的安全性

    Institute of Scientific and Technical Information of China (English)

    章浩; 叶真龙; 钱其军

    2016-01-01

    Objective To investigate whether decreasing affinity of CAR-T cells can increase their therapeutic outcome or not .Methods Moderate affinity La-G3HER2-CAR and high affinity Ha-G3HER2-CAR were constructed ,and electroporated to modify T cells .Western blot assay ,FCM assay and the RTCA DP cytotoxic equipment were applied to test the CAR expres-sion and cytotoxic function of CAR-T cells .Results 43000 and 58000 exogenous CD3ζfragments were expressed by both La-G3HER2-CAR-T cells and Ha-G3HER2-CAR-T cells with 58 .1% and 69 .0% transfection rate respectively .High affinity Ha-G3HER2-CAR-T cells effectively killed all target tumor cells by which HER2 was expressed at variable expression levels , while moderate affinity La-G3HER2-CAR-T cells specifically killed HER2 high-level expressing SK-OV-3 and BT474 cells ,and showed weaker cytotoxicity on HER2 moderate-level expressing MDA-MB-231 and HCC-202 cells ,and showed no cytotoxicity on HER2 low-level expressing MCF-7 and 293 cells .The underlying mechanic investigation found that La-G3HER2-CAR-T cells and Ha-G3HER2-CAR-T cells were differentially activated by co-culture with MDA-MB-231 (CD107a:8 .2% vs 71 .6% , IFN-γ:66 .3% vs 83 .4% ,TNF-α:73 .4% vs 94 .1% ) .Conclusion Moderate affinity La-G3HER2-CAR-T cells have en-hanced specific cytotoxicity toward target tumor cells compared to high affinity Ha-G3HER2-CAR-T cells ,decreasing affinity of CAR-T cell is a promising strategy to increase the therapeutic outcome of CAR-T cell based immunotherapies .%目的 探讨降低CAR-T细胞亲和力是否能有效提高其杀伤特异性,减少"脱靶效应".方法 构建靶向HER2的中等亲和力和高亲和力的La-G3HER2-CAR和Ha-G3HER2-CAR并电穿孔转染T细胞,采用Western Blot、FCM技术和xCELLigence RTCA DP进行CAR载体表达和杀伤功能检测.结果 La-G3HER2-CAR-T细胞和Ha-G3HER2-CAR-T细胞分别表达43000和58000的外源CAR载体片段,转染效率分别为58.1%和69.0%.高亲和力的Ha-G3HER2-CAR-T

  5. Targeting B cell responses in universal influenza vaccine design

    Science.gov (United States)

    Kaur, Kaval; Sullivan, Meghan; Wilson, Patrick C

    2011-01-01

    Since its first administration in the 1940s, the influenza vaccine has provided tremendous relief against influenza infections. However, time has revealed the vaccine’s ultimate limit and the call for its reinvention has now come, just as we are beginning to appreciate the antibody immune responses vital in preventing infections. New strategies to design the influenza vaccine rely on selectively inducing broadly neutralizing antibodies that are specific for highly conserved viral epitopes. Such approaches take us away from the limited range of protection provided by current seasonal influenza vaccines and towards a future with a pan-influenza vaccine capable of providing universal strain coverage. PMID:21940217

  6. Structure-function studies of DNA binding domain of response regulator KdpE reveals equal affinity interactions at DNA half-sites.

    Directory of Open Access Journals (Sweden)

    Anoop Narayanan

    Full Text Available Expression of KdpFABC, a K(+ pump that restores osmotic balance, is controlled by binding of the response regulator KdpE to a specific DNA sequence (kdpFABC(BS via the winged helix-turn-helix type DNA binding domain (KdpE(DBD. Exploration of E. coli KdpE(DBD and kdpFABC(BS interaction resulted in the identification of two conserved, AT-rich 6 bp direct repeats that form half-sites. Despite binding to these half-sites, KdpE(DBD was incapable of promoting gene expression in vivo. Structure-function studies guided by our 2.5 Å X-ray structure of KdpE(DBD revealed the importance of residues R193 and R200 in the α-8 DNA recognition helix and T215 in the wing region for DNA binding. Mutation of these residues renders KdpE incapable of inducing expression of the kdpFABC operon. Detailed biophysical analysis of interactions using analytical ultracentrifugation revealed a 2∶1 stoichiometry of protein to DNA with dissociation constants of 200±100 and 350±100 nM at half-sites. Inactivation of one half-site does not influence binding at the other, indicating that KdpE(DBD binds independently to the half-sites with approximately equal affinity and no discernable cooperativity. To our knowledge, these data are the first to describe in quantitative terms the binding at half-sites under equilibrium conditions for a member of the ubiquitous OmpR/PhoB family of proteins.

  7. "Siglec"ting the allergic response for therapeutic targeting.

    Science.gov (United States)

    Bochner, Bruce S

    2016-06-01

    As a physician-scientist, I have pursued research related to translational immunology with the goal of improving our ability to diagnose and treat allergic, immunologic and other diseases involving eosinophils, basophils and mast cells. We have tried to delineate novel mechanisms of human disease, working whenever possible with primary human cells and tissues, attempting to identify targets that might be amenable to the development of new therapies. As a general strategy, we have compared eosinophils, basophils, mast cells and neutrophils to look for pathways in inflammation that were unique to distinct subsets of these cells. In doing so, the concepts of glycobiology did not enter my mind until we began noticing some intriguing functional differences involving selectins and their ligands among these cell types. One simple observation, that neutrophils were coated with a glycan that allowed them to interact with an endothelial adhesion molecule while eosinophils lacked this structure, pried open the glyco-door for me. Fruitful collaborations with card-carrying glycobiologists soon followed that have forever positively influenced our science, and have enhanced our hypotheses, experimental design, research opportunities and discoveries. Within a few years, we helped to discover Siglec-8, an I-type lectin expressed only on human eosinophils, basophils, mast cells. This receptor, together with its closest mouse counterpart Siglec-F, has been the primary focus of our work now for over a decade. If not for those in the fields of glycobiology and glycoimmunology, my lab would not have made much progress toward the goal of leveraging Siglec-8 for therapeutic purposes. PMID:26911285

  8. Porous Matrix Stiffness Modulates Response to Targeted Therapy in Breast Carcinoma.

    Science.gov (United States)

    Liu, Cuiying; Li, Xiang; Hua, Wenda; Li, Jianjun; Han, Xinxiao; Ha, Qing; Feng, Jiantao; Liao, Fulong; Li, Dongguo; Han, Dong

    2016-09-01

    Porous matrix stiffness modulates response to targeted therapy. Poroelastic behavior within porous matrix may modulate the molecule events in cell-matrix and cell-cell interaction like the complex formation of human epidermal growth factor receptor-2 (HER2)-Src-α6β4 integrin, influencing the targeted therapy with lapatinib.

  9. MPLA incorporation into DC-targeting glycoliposomes favours anti-tumour T cell responses

    NARCIS (Netherlands)

    Boks, Martine A.; Ambrosini, Martino; Bruijns, Sven C.; Kalay, Hakan; Van Bloois, Louis; Storm, G; Garcia-Vallejo, Juan J.; Van Kooyk, Yvette

    2015-01-01

    Abstract Dendritic cells (DC) are attractive targets for cancer immunotherapy as they initiate strong and long-lived tumour-specific T cell responses. DC can be effectively targeted in vivo with tumour antigens by using nanocarriers such as liposomes. Cross-presentation of tumour antigens is enhance

  10. MPLA incorporation into DC-targeting glycoliposomes favours anti-tumour T cell responses

    NARCIS (Netherlands)

    Boks, M.A.; Ambrosini, Martino; Bruijns, Sven C.M.; Kalay, Hakan; Bloois, van Louis; Storm, G.; Garcia-Vallejo, Juan J.; Kooyk, van Y.

    2015-01-01

    Dendritic cells (DC) are attractive targets for cancer immunotherapy as they initiate strong and long-lived tumour-specific T cell responses. DC can be effectively targeted in vivo with tumour antigens by using nanocarriers such as liposomes. Cross-presentation of tumour antigens is enhanced with st

  11. Affinity Proteomics in the mountains: Alpbach 2015.

    Science.gov (United States)

    Taussig, Michael J

    2016-09-25

    The 2015 Alpbach Workshop on Affinity Proteomics, organised by the EU AFFINOMICS consortium, was the 7th workshop in this series. As in previous years, the focus of the event was the current state of affinity methods for proteome analysis, including complementarity with mass spectrometry, progress in recombinant binder production methods, alternatives to classical antibodies as affinity reagents, analysis of proteome targets, industry focus on biomarkers, and diagnostic and clinical applications. The combination of excellent science with Austrian mountain scenery and winter sports engender an atmosphere that makes this series of workshops exceptional. The articles in this Special Issue represent a cross-section of the presentations at the 2015 meeting. PMID:27118167

  12. Affinity-Based Screening of Tetravalent Peptides Identifies Subtype-Selective Neutralizers of Shiga Toxin 2d, a Highly Virulent Subtype, by Targeting a Unique Amino Acid Involved in Its Receptor Recognition.

    Science.gov (United States)

    Mitsui, Takaaki; Watanabe-Takahashi, Miho; Shimizu, Eiko; Zhang, Baihao; Funamoto, Satoru; Yamasaki, Shinji; Nishikawa, Kiyotaka

    2016-09-01

    Shiga toxin (Stx), a major virulence factor of enterohemorrhagic Escherichia coli (EHEC), can be classified into two subgroups, Stx1 and Stx2, each consisting of various closely related subtypes. Stx2 subtypes Stx2a and Stx2d are highly virulent and linked with serious human disorders, such as acute encephalopathy and hemolytic-uremic syndrome. Through affinity-based screening of a tetravalent peptide library, we previously developed peptide neutralizers of Stx2a in which the structure was optimized to bind to the B-subunit pentamer. In this study, we identified Stx2d-selective neutralizers by targeting Asn16 of the B subunit, an amino acid unique to Stx2d that plays an essential role in receptor binding. We synthesized a series of tetravalent peptides on a cellulose membrane in which the core structure was exactly the same as that of peptides in the tetravalent library. A total of nine candidate motifs were selected to synthesize tetravalent forms of the peptides by screening two series of the tetravalent peptides. Five of the tetravalent peptides effectively inhibited the cytotoxicity of Stx2a and Stx2d, and notably, two of the peptides selectively inhibited Stx2d. These two tetravalent peptides bound to the Stx2d B subunit with high affinity dependent on Asn16. The mechanism of binding to the Stx2d B subunit differed from that of binding to Stx2a in that the peptides covered a relatively wide region of the receptor-binding surface. Thus, this highly optimized screening technique enables the development of subtype-selective neutralizers, which may lead to more sophisticated treatments of infections by Stx-producing EHEC. PMID:27382021

  13. The specificity of targeted vaccines for APC surface molecules influences the immune response phenotype.

    Directory of Open Access Journals (Sweden)

    Gunnveig Grødeland

    Full Text Available Different diseases require different immune responses for efficient protection. Thus, prophylactic vaccines should prime the immune system for the particular type of response needed for protection against a given infectious agent. We have here tested fusion DNA vaccines which encode proteins that bivalently target influenza hemagglutinins (HA to different surface molecules on antigen presenting cells (APC. We demonstrate that targeting to MHC class II molecules predominantly induced an antibody/Th2 response, whereas targeting to CCR1/3/5 predominantly induced a CD8(+/Th1 T cell response. With respect to antibodies, the polarizing effect was even more pronounced upon intramuscular (i.m delivery as compared to intradermal (i.d. vaccination. Despite these differences in induced immune responses, both vaccines protected against a viral challenge with influenza H1N1. Substitution of HA with ovalbumin (OVA demonstrated that polarization of immune responses, as a consequence of APC targeting specificity, could be extended to other antigens. Taken together, the results demonstrate that vaccination can be tailor-made to induce a particular phenotype of adaptive immune responses by specifically targeting different surface molecules on APCs.

  14. Rational development of high-affinity T-cell receptor-like antibodies.

    Science.gov (United States)

    Stewart-Jones, Guillaume; Wadle, Andreas; Hombach, Anja; Shenderov, Eugene; Held, Gerhard; Fischer, Eliane; Kleber, Sascha; Nuber, Natko; Stenner-Liewen, Frank; Bauer, Stefan; McMichael, Andrew; Knuth, Alexander; Abken, Hinrich; Hombach, Andreas A; Cerundolo, Vincenzo; Jones, E Yvonne; Renner, Christoph

    2009-04-01

    T-cell interaction with a target cell is a key event in the adaptive immune response and primarily driven by T-cell receptor (TCR) recognition of peptide-MHC (pMHC) complexes. TCR avidity for a given pMHC is determined by number of MHC molecules, availability of coreceptors, and TCR affinity for MHC or peptide, respectively, with peptide recognition being the most important factor to confer target specificity. Here we present high-resolution crystal structures of 2 Fab antibodies in complex with the immunodominant NY-ESO-1(157-165) peptide analogue (SLLMWITQV) presented by HLA-A*0201 and compare them with a TCR recognizing the same pMHC. Binding to the central methionine-tryptophan peptide motif and orientation of binding were almost identical for Fabs and TCR. As the MW "peg" dominates the contacts between Fab and peptide, we estimated the contributions of individual amino acids between the Fab and peptide to provide the rational basis for a peptide-focused second-generation, high-affinity antibody library. The final Fab candidate achieved better peptide binding by 2 light-chain mutations, giving a 20-fold affinity improvement to 2-4 nM, exceeding the affinity of the TCR by 1,000-fold. The high-affinity Fab when grafted as recombinant TCR on T cells conferred specific killing of HLA-A*0201/NY-ESO-1(157-165) target cells. In summary, we prove that affinity maturation of antibodies mimicking a TCR is possible and provide a strategy for engineering high-affinity antibodies that can be used in targeting specific pMHC complexes for diagnostic and therapeutic purposes. PMID:19307587

  15. In vitro and in vivo evaluation of a (18F-labeled high affinity NOTA conjugated bombesin antagonist as a PET ligand for GRPR-targeted tumor imaging.

    Directory of Open Access Journals (Sweden)

    Zohreh Varasteh

    Full Text Available Expression of the gastrin-releasing peptide receptor (GRPR in prostate cancer suggests that this receptor can be used as a potential molecular target to visualize and treat these tumors. We have previously investigated an antagonist analog of bombesin (D-Phe-Gln-Trp-Ala-Val-Gly-His-Sta-Leu-NH2, RM26 conjugated to 1,4,7-triazacyclononane-N,N',N''-triacetic acid (NOTA via a diethylene glycol (PEG2 spacer (NOTA-P2-RM26 labeled with (68Ga and (111In. We found that this conjugate has favorable properties for in vivo imaging of GRPR-expression. The focus of this study was to develop a (18F-labelled PET agent to visualize GRPR. NOTA-P2-RM26 was labeled with (18F using aluminum-fluoride chelation. Stability, in vitro binding specificity and cellular processing tests were performed. The inhibition efficiency (IC50 of the [(natF]AlF-NOTA-P2-RM26 was compared to that of the (natGa-loaded peptide using (125I-Tyr(4-BBN as the displacement radioligand. The pharmacokinetics and in vivo binding specificity of the compound were studied. NOTA-P2-RM26 was labeled with (18F within 1 h (60-65% decay corrected radiochemical yield, 55 GBq/µmol. The radiopeptide was stable in murine serum and showed high specific binding to PC-3 cells. [(natF]AlF-NOTA-P2-RM26 showed a low nanomolar inhibition efficiency (IC50=4.4±0.8 nM. The internalization rate of the tracer was low. Less than 14% of the cell-bound radioactivity was internalized after 4 h. The biodistribution of [(18F]AlF-NOTA-P2-RM26 demonstrated rapid blood clearance, low liver uptake and low kidney retention. The tumor uptake at 3 h p.i. was 5.5±0.7 %ID/g, and the tumor-to-blood, -muscle and -bone ratios were 87±42, 159±47, 38±16, respectively. The uptake in tumors, pancreas and other GRPR-expressing organs was significantly reduced when excess amount of non-labeled peptide was co-injected. The low uptake in bone suggests a high in vivo stability of the Al-F bond. High contrast PET image was obtained 3 h p

  16. Electromagnetic modelling of Ground Penetrating Radar responses to complex targets

    Science.gov (United States)

    Pajewski, Lara; Giannopoulos, Antonis

    2014-05-01

    defined through a constant real value, or else its frequency-dispersion properties can be taken into account by incorporating into the model Debye approximations. The electromagnetic source can be represented as a simple line of current (in the case of two-dimensional models), a Hertzian dipole, a bow tie antenna, or else, the realistic description of a commercial antenna can be included in the model [2]. Preliminary results for some of the proposed cells are presented, obtained by using GprMax [3], a freeware tool which solves Maxwell's equations by using a second order in space and time Finite-Difference Time-Domain algorithm. B-Scans and A-Scans are calculated at 1.5 GHz, for the total electric field and for the field back-scattered by targets embedded in the cells. A detailed description of the structures, together with the relevant numerical results obtained to date, are available for the scientific community on the website of COST Action TU1208, www.GPRadar.eu. Research groups working on the development of electromagnetic forward- and inverse-scattering techniques, as well as on imaging methods, might test and compare the accuracy and applicability of their approaches on the proposed set of scenarios. The aim of this initiative is not that of identifying the best methods, but more properly to indicate the range of reliability of each approach, highlighting its advantages and drawbacks. In the future, the realisation of the proposed concrete cells and the acquisition of GPR experimental data would allow a very effective benchmark for forward and inverse scattering methods. References [1] R. Yelf, A. Ward, "Nine steps to concrete wisdom." Proc. 13th International Conference on Ground Penetrating Radar, Lecce, Italy, 21-25 June 2010, pp. 1-8. [2] C. Warren, A. Giannopoulos, "Creating FDTD models of commercial GPR antennas using Taguchi's optimisation method." Geophysics (2011), 76, article ID G37. [3] A. Giannopoulos, "Modelling ground penetrating radar by GPRMAX

  17. Artificial ground motion compatible with specified ground shaking peaks and target response spectrum

    Institute of Scientific and Technical Information of China (English)

    Zhao Fengxin; Zhang Yushan; Lü Hongshan

    2006-01-01

    This article describes a hybrid simulation method to generate artificial ground motion time histories that are compatible with specified peak seismic acceleration, velocity and displacement as well as the target response spectrum of absolute acceleration. First, based on traditional methods that match the target spectrum in the frequency domain, an initial acceleration time history was synthesized to satisfy the specified peak acceleration, target spectral acceleration and intensity envelope. Second, by using the inversion formula of the seismic input to a linear single-degree-of-freedom system and by superimposing a series of narrow-band time histories in the time domain, the initial time history is further modified to allow its peak velocity and displacement to approach the targets and improve its matching precision with the target spectrum.Numerical examples are provided to demonstrate that the proposed method achieves good agreement with the target values.

  18. High-affinity human leucocyte antigen class I binding variola-derived peptides induce CD4(+) T cell responses more than 30 years post-vaccinia virus vaccination

    DEFF Research Database (Denmark)

    Wang, M.; Tang, Sheila Tuyet; Lund, Ole;

    2009-01-01

    Interferon-gamma secreting T lymphocytes against pox virus-derived synthetic 9-mer peptides were tested by enzyme-linked immunospot in peripheral blood of individuals vaccinated with vaccinia virus more than 30 years ago. The peptides were characterized biochemically as high-affinity human...

  19. Affine dynamics with torsion

    Energy Technology Data Exchange (ETDEWEB)

    Gueltekin, Kemal [Izmir Institute of Technology, Department of Physics, Izmir (Turkey)

    2016-03-15

    In this study, we give a thorough analysis of a general affine gravity with torsion. After a brief exposition of the affine gravities considered by Eddington and Schroedinger, we construct and analyze different affine gravities based on the determinants of the Ricci tensor, the torsion tensor, the Riemann tensor, and their combinations. In each case we reduce equations of motion to their simplest forms and give a detailed analysis of their solutions. Our analyses lead to the construction of the affine connection in terms of the curvature and torsion tensors. Our solutions of the dynamical equations show that the curvature tensors at different points are correlated via non-local, exponential rescaling factors determined by the torsion tensor. (orig.)

  20. Characterization of wheat miRNAs and their target genes responsive to cadmium stress.

    Science.gov (United States)

    Qiu, ZongBo; Hai, BenZhai; Guo, JunLi; Li, YongFang; Zhang, Liang

    2016-04-01

    A increasing number of microRNAs have been shown to play important regulatory roles in plant responses to various metal stresses. However, little information about miRNAs especially miRNAs responsive to cadmium (Cd) stress is available in wheat. To investigate the role of miRNAs in responses to Cd stress, wheat seedlings were subjected to 250 μM Cd solution for 6, 12, 24 and 48 h, and analyses of morphological and physiological changes as well as the expression of five miRNAs and their corresponding targets were carried out. Our results demonstrated that miRNAs and their targets were differentially expressed in leaves and roots of wheat seedlings exposed to Cd stress. Furthermore, miR398 may involve in oxidative stress tolerance by regulating its target CSD to participate in Cd stress. Among ten miRNA-target pairs studied, nine pairs showed complex regulation relationship in leaves and roots of wheat seedlings exposed to Cd stress. These findings suggested that miRNAs are involved in the mediation of Cd stress signaling responses in wheat. The characterization of the miRNAs and the associated targets in responses to Cd exposure provides a framework for understanding the molecular mechanism of heavy metal tolerance in plants. PMID:26854408

  1. Affine and degenerate affine BMW algebras: Actions on tensor space

    CERN Document Server

    Daugherty, Zajj; Virk, Rahbar

    2012-01-01

    The affine and degenerate affine Birman-Murakami-Wenzl (BMW) algebras arise naturally in the context of Schur-Weyl duality for orthogonal and symplectic quantum groups and Lie algebras, respectively. Cyclotomic BMW algebras, affine and cyclotomic Hecke algebras, and their degenerate versions are quotients. In this paper we explain how the affine and degenerate affine BMW algebras are tantalizers (tensor power centralizer algebras) by defining actions of the affine braid group and the degenerate affine braid algebra on tensor space and showing that, in important cases, these actions induce actions of the affine and degenerate affine BMW algebras. We then exploit the connection to quantum groups and Lie algebras to determine universal parameters for the affine and degenerate affine BMW algebras. Finally, we show that the universal parameters are central elements--the higher Casimir elements for orthogonal and symplectic enveloping algebras and quantum groups.

  2. Doing worse, but feeling happy: Social comparison and identification in response to upward and downward targets

    OpenAIRE

    Groothof, H.A.K.; Siero, F.W.; Buunk, A.P.

    2008-01-01

    We investigated people’s responses to exposure to downward and upward targets. In Study 1, among 197 participants, it was predicted and found that such exposure led to a contrast effect on self-evaluation, and to an assimilation effect on affect. In Study 2, among 148 participants, it was predicted and found that the contrast effect on self-evaluation occurred in particular when participants were induced to compare themselves with the target, and that the assimilation effect on affect occurre...

  3. Seed coat color and seed weight contribute differential responses of targeted metabolites in soybean seeds.

    Science.gov (United States)

    Lee, Jinwook; Hwang, Young-Sun; Kim, Sun Tae; Yoon, Won-Byong; Han, Won Young; Kang, In-Kyu; Choung, Myoung-Gun

    2017-01-01

    The distribution and variation of targeted metabolites in soybean seeds are affected by genetic and environmental factors. In this study, we used 192 soybean germplasm accessions collected from two provinces of Korea to elucidate the effects of seed coat color and seeds dry weight on the metabolic variation and responses of targeted metabolites. The effects of seed coat color and seeds dry weight were present in sucrose, total oligosaccharides, total carbohydrates and all measured fatty acids. The targeted metabolites were clustered within three groups. These metabolites were not only differently related to seeds dry weight, but also responded differentially to seed coat color. The inter-relationship between the targeted metabolites was highly present in the result of correlation analysis. Overall, results revealed that the targeted metabolites were diverged in relation to seed coat color and seeds dry weight within locally collected soybean seed germplasm accessions. PMID:27507473

  4. Ablative response of microdisc targets irradiated by 1.064 μm light

    International Nuclear Information System (INIS)

    The ablative response of microdisc targets irradiated with an 8 ns FWHM Nd:Glass laser is compared to that of infinite foils. The front surface exhaust velocity, plasma electron temperature, and size of the x-ray emitting region of Al and Fe targets irradiated with a laser intensity of 1013 to 1014 W/cm2 are relatively independent of the target diameter and thickness, but the x-ray conversion efficiency (E/sub photon/ greater than or equal to 700 eV) decreases as these target dimensions decrease. These observations suggest that in this experiment: (1) lateral heat flow is not a significant factor: (2) a steady-state ablative flow at the front surface is established; and (3) two-dimensional effects lead to an increase in the density gradient or to a decrease in the average density in the vicinity of the x-ray emitting region as the target dimensions decrease

  5. Individualized therapies in colorectal cancer: KRAS as a marker for response to EGFR-targeted therapy

    Directory of Open Access Journals (Sweden)

    Li Kuiyuan

    2009-04-01

    Full Text Available Abstract Individualized therapies that are tailored to a patient's genetic composition will be of tremendous value for treatment of cancer. Recently, Kirsten ras (KRAS status has emerged as a predictor of response to epidermal growth factor receptor (EGFR targeted therapies. In this article, we will discuss targeted therapies for colorectal cancers (CRC based on EGFR signaling pathway and review published data about the potential usefulness of KRAS as a biological marker for response to these therapies. Results from relevant studies published since 2005 and unpublished results presented at national meetings were retrieved and summarized. These studies reflected response (or lack of response to EGFR-targeted therapies in patients with metastatic CRC as a function of KRAS status. It has become clear that patients with colorectal cancer whose tumor has an activating mutation in KRAS do not respond to monoclonal antibody therapies targeting EGFR. It should now become a standard practice that any patients being considered for EGFR targeted therapies have their tumors tested for KRAS status and only those with wild-type KRAS being offered such therapies.

  6. Application of Adenosine Triphosphate Affinity Probe and Scheduled Multiple-Reaction Monitoring Analysis for Profiling Global Kinome in Human Cells in Response to Arsenite Treatment

    OpenAIRE

    Guo, Lei; Xiao, Yongsheng; Wang, Yinsheng

    2014-01-01

    Phosphorylation of cellular components catalyzed by kinases plays important roles in cell signaling and proliferation. Quantitative assessment of perturbation in global kinome may provide crucial knowledge for elucidating the mechanisms underlying the cytotoxic effects of environmental toxicants. Here, we utilized an adenosine triphosphate (ATP) affinity probe coupled with stable isotope labeling by amino acids in cell culture (SILAC) to assess quantitatively the arsenite-induced alteration o...

  7. Competitive Selection from Single Domain Antibody Libraries Allows Isolation of High-Affinity Antihapten Antibodies That Are Not Favored in the llama Immune Response

    OpenAIRE

    Rosa, Sofia Tabares-da; Rossotti, Martin; Carleiza, Carmen; Carrión, Federico; Pritsch, Otto; Ahn, Ki Chang; Last, Jerold A.; Hammock, Bruce D.; González-Sapienza, Gualberto

    2011-01-01

    Single-domain antibodies (sdAbs) found in camelids, lack a light chain and their antigen-binding site sits completely in the heavy-chain variable domain (VHH). Their simplicity, thermostability, and ease in expression have made VHHs highly attractive. While this has been successfully exploited for macromolecular antigens, their application to the detection of small molecules is still limited to a very few reports, mostly describing low affinity VHHs. Using triclocarban (TCC) as a model hapten...

  8. Impact velocity vs. target hardness relationships for equivalent response of cask structures

    International Nuclear Information System (INIS)

    In this paper, impact velocity vs. target hardness relationships for cask structures are reviewed. The relationships are based on equivalent cask responses in terms of equal deceleration or similar cask damages. By examining several past cask or container tests as well as some analytical results, some conclusions can be drawn about the relationship between target hardness and equivalent impact velocities. This relationship clearly shows that the cask response to impact is cask-dependent and that the rigid sphere impact model results in an unconservative estimate of equivalent velocity

  9. Balancing search and target response in cooperative unmanned aerial vehicle (UAV) teams.

    Science.gov (United States)

    Jin, Yan; Liao, Yan; Minai, Ali A; Polycarpou, Marios M

    2006-06-01

    This paper considers a heterogeneous team of cooperating unmanned aerial vehicles (UAVs) drawn from several distinct classes and engaged in a search and action mission over a spatially extended battlefield with targets of several types. During the mission, the UAVs seek to confirm and verifiably destroy suspected targets and discover, confirm, and verifiably destroy unknown targets. The locations of some (or all) targets are unknown a priori, requiring them to be located using cooperative search. In addition, the tasks to be performed at each target location by the team of cooperative UAVs need to be coordinated. The tasks must, therefore, be allocated to UAVs in real time as they arise, while ensuring that appropriate vehicles are assigned to each task. Each class of UAVs has its own sensing and attack capabilities, so the need for appropriate assignment is paramount. In this paper, an extensive dynamic model that captures the stochastic nature of the cooperative search and task assignment problems is developed, and algorithms for achieving a high level of performance are designed. The paper focuses on investigating the value of predictive task assignment as a function of the number of unknown targets and number of UAVs. In particular, it is shown that there is a tradeoff between search and task response in the context of prediction. Based on the results, a hybrid algorithm for switching the use of prediction is proposed, which balances the search and task response. The performance of the proposed algorithms is evaluated through Monte Carlo simulations.

  10. Response of Solid and Liquid Targets to High Power Proton Beams for Neutrino Factories

    CERN Document Server

    Sievers, P

    2000-01-01

    The response of solid and liquid targets to rapid heating by the incident proton beam is assessed in a classical way, among other things by solving the wave equation under linear conditions and in cylindrical symmetry. This study provides bench mark values and allows to identify critical issues and limiting factors which can help to guide further investigations with more sophisticated means.

  11. Differential inflammatory response to inhaled lipopolysaccharide targeted either to the airways or the alveoli in man.

    Directory of Open Access Journals (Sweden)

    Winfried Möller

    Full Text Available Endotoxin (Lipopolysaccharide, LPS is a potent inducer of inflammation and there is various LPS contamination in the environment, being a trigger of lung diseases and exacerbation. The objective of this study was to assess the time course of inflammation and the sensitivities of the airways and alveoli to targeted LPS inhalation in order to understand the role of LPS challenge in airway disease.In healthy volunteers without any bronchial hyperresponsiveness we targeted sequentially 1, 5 and 20 µg LPS to the airways and 5 µg LPS to the alveoli using controlled aerosol bolus inhalation. Inflammatory parameters were assessed during a 72 h time period. LPS deposited in the airways induced dose dependent systemic responses with increases of blood neutrophils (peaking at 6 h, Interleukin-6 (peaking at 6 h, body temperature (peaking at 12 h, and CRP (peaking at 24 h. 5 µg LPS targeted to the alveoli caused significantly stronger effects compared to 5 µg airway LPS deposition. Local responses were studied by measuring lung function (FEV(1 and reactive oxygen production, assessed by hydrogen peroxide (H(2O(2 in fractionated exhaled breath condensate (EBC. FEV(1 showed a dose dependent decline, with lowest values at 12 h post LPS challenge. There was a significant 2-fold H(2O(2 induction in airway-EBC at 2 h post LPS inhalation. Alveolar LPS targeting resulted in the induction of very low levels of EBC-H(2O(2.Targeting LPS to the alveoli leads to stronger systemic responses compared to airway LPS targeting. Targeted LPS inhalation may provide a novel model of airway inflammation for studying the role of LPS contamination of air pollution in lung diseases, exacerbation and anti-inflammatory drugs.

  12. Direct binding targets of the stringent response alarmone (p)ppGpp.

    Science.gov (United States)

    Kanjee, Usheer; Ogata, Koji; Houry, Walid A

    2012-09-01

    The Escherichia coli stringent response, mediated by the alarmone ppGpp, is responsible for the reorganization of cellular transcription upon nutritional starvation and other stresses. These transcriptional changes occur mainly as a result of the direct effects of ppGpp and its partner transcription factor DksA on RNA polymerase. An often overlooked feature of the stringent response is the direct targeting of other proteins by ppGpp. Here we review the literature on proteins that are known to bind ppGpp and, based on sequence homology, X-ray crystal structures and in silico docking, we propose new potential protein binding targets for ppGpp. These proteins were found to fall into five main categories: (i) cellular GTPases, (ii) proteins involved in nucleotide metabolism, (iii) proteins involved in lipid metabolism, (iv) general metabolic proteins and (v) PLP-dependent basic aliphatic amino acid decarboxylases. Bioinformatic rationale is provided for expanding the role of ppGpp in regulating the activities of the cellular GTPases. Proteins involved in nucleotide and lipid metabolism and general metabolic proteins provide an interesting set of structurally varied stringent response targets. While the inhibition of some PLP-dependent decarboxylases by ppGpp suggests the existence of cross-talk between the acid stress and stringent response systems.

  13. Affine stochastic mortality

    NARCIS (Netherlands)

    D.F. Schrager

    2006-01-01

    We propose a new model for stochastic mortality. The model is based on the literature on affine term structure models. It satisfies three important requirements for application in practice: analytical tractibility, clear interpretation of the factors and compatibility with financial option pricing m

  14. CERN antiproton target: Hydrocode analysis of its core material dynamic response under proton beam impact

    Science.gov (United States)

    Martin, Claudio Torregrosa; Perillo-Marcone, Antonio; Calviani, Marco; Muñoz-Cobo, José-Luis

    2016-07-01

    Antiprotons are produced at CERN by colliding a 26 GeV /c proton beam with a fixed target made of a 3 mm diameter, 55 mm length iridium core. The inherent characteristics of antiproton production involve extremely high energy depositions inside the target when impacted by each primary proton beam, making it one of the most dynamically demanding among high energy solid targets in the world, with a rise temperature above 2000 °C after each pulse impact and successive dynamic pressure waves of the order of GPa's. An optimized redesign of the current target is foreseen for the next 20 years of operation. As a first step in the design procedure, this numerical study delves into the fundamental phenomena present in the target material core under proton pulse impact and subsequent pressure wave propagation by the use of hydrocodes. Three major phenomena have been identified, (i) the dominance of a high frequency radial wave which produces destructive compressive-to-tensile pressure response (ii) The existence of end-of-pulse tensile waves and its relevance on the overall response (iii) A reduction of 44% in tensile pressure could be obtained by the use of a high density tantalum cladding.

  15. From drug response profiling to target addiction scoring in cancer cell models

    Directory of Open Access Journals (Sweden)

    Bhagwan Yadav

    2015-10-01

    Full Text Available Deconvoluting the molecular target signals behind observed drug response phenotypes is an important part of phenotype-based drug discovery and repurposing efforts. We demonstrate here how our network-based deconvolution approach, named target addiction score (TAS, provides insights into the functional importance of druggable protein targets in cell-based drug sensitivity testing experiments. Using cancer cell line profiling data sets, we constructed a functional classification across 107 cancer cell models, based on their common and unique target addiction signatures. The pan-cancer addiction correlations could not be explained by the tissue of origin, and only correlated in part with molecular and genomic signatures of the heterogeneous cancer cells. The TAS-based cancer cell classification was also shown to be robust to drug response data resampling, as well as predictive of the transcriptomic patterns in an independent set of cancer cells that shared similar addiction signatures with the 107 cancers. The critical protein targets identified by the integrated approach were also shown to have clinically relevant mutation frequencies in patients with various cancer subtypes, including not only well-established pan-cancer genes, such as PTEN tumor suppressor, but also a number of targets that are less frequently mutated in specific cancer types, including ABL1 oncoprotein in acute myeloid leukemia. An application to leukemia patient primary cell models demonstrated how the target deconvolution approach offers functional insights into patient-specific addiction patterns, such as those indicative of their receptor-type tyrosine-protein kinase FLT3 internal tandem duplication (FLT3-ITD status and co-addiction partners, which may lead to clinically actionable, personalized drug treatment developments. To promote its application to the future drug testing studies, we have made available an open-source implementation of the TAS calculation in the form

  16. Rapid antibody responses by low-dose, single-step, dendritic cell-targeted immunization

    OpenAIRE

    Wang, Hui; Griffiths, Michelle N.; Burton, Dennis R; Ghazal, Peter

    2000-01-01

    We have compared the kinetics of antibody responses in conventional and dendritic cell-targeted immunization by using a model antigen in mice. Targeting was achieved by linking the reporter antigen (polyclonal goat anti-hamster antibody) to N418, a hamster mAb that binds to the CD11c molecule on the surface of murine dendritic cells. Intradermal injection of submicrogram quantities of goat anti-hamster antibody complexed to mAb N418 elicited goat antibody-specific serum IgG in mice. Antigen-s...

  17. Generating Targeted Gene Knockout Lines in Physcomitrella patens to Study Evolution of Stress-Responsive Mechanisms

    Science.gov (United States)

    Maronova, Monika; Kalyna, Maria

    2016-01-01

    The moss Physcomitrella patens possesses highly efficient homologous recombination allowing targeted gene manipulations and displays many features of the early land plants including high tolerance to abiotic stresses. It is therefore an invaluable model organism for studies of gene functions and comparative studies of evolution of stress responses in plants. Here, we describe a method for generating targeted gene knockout lines in P. patens using a polyethylene glycol-mediated transformation of protoplasts including basic in vitro growth, propagation, and maintenance techniques. PMID:26867627

  18. RNF111/Arkadia is a SUMO-targeted ubiquitin ligase that facilitates the DNA damage response

    DEFF Research Database (Denmark)

    Poulsen, Sara L; Hansen, Rebecca K; Wagner, Sebastian A;

    2013-01-01

    Protein modifications by ubiquitin and small ubiquitin-like modifier (SUMO) play key roles in cellular signaling pathways. SUMO-targeted ubiquitin ligases (STUbLs) directly couple these modifications by selectively recognizing SUMOylated target proteins through SUMO-interacting motifs (SIMs......)-induced SUMOylation and ubiquitylation. Moreover, we show that RNF111 facilitated NER by regulating the recruitment of XPC to UV-damaged DNA. Our findings establish RNF111 as a new STUbL that directly links nonproteolytic ubiquitylation and SUMOylation in the DNA damage response....

  19. Memory T cell responses targeting the SARS coronavirus persist up to 11 years post-infection.

    Science.gov (United States)

    Ng, Oi-Wing; Chia, Adeline; Tan, Anthony T; Jadi, Ramesh S; Leong, Hoe Nam; Bertoletti, Antonio; Tan, Yee-Joo

    2016-04-12

    Severe acute respiratory syndrome (SARS) is a highly contagious infectious disease which first emerged in late 2002, caused by a then novel human coronavirus, SARS coronavirus (SARS-CoV). The virus is believed to have originated from bats and transmitted to human through intermediate animals such as civet cats. The re-emergence of SARS-CoV remains a valid concern due to the continual persistence of zoonotic SARS-CoVs and SARS-like CoVs (SL-CoVs) in bat reservoirs. In this study, the screening for the presence of SARS-specific T cells in a cohort of three SARS-recovered individuals at 9 and 11 years post-infection was carried out, and all memory T cell responses detected target the SARS-CoV structural proteins. Two CD8(+) T cell responses targeting the SARS-CoV membrane (M) and nucleocapsid (N) proteins were characterized by determining their HLA restriction and minimal T cell epitope regions. Furthermore, these responses were found to persist up to 11 years post-infection. An absence of cross-reactivity of these CD8(+) T cell responses against the newly-emerged Middle East respiratory syndrome coronavirus (MERS-CoV) was also demonstrated. The knowledge of the persistence of SARS-specific celullar immunity targeting the viral structural proteins in SARS-recovered individuals is important in the design and development of SARS vaccines, which are currently unavailable. PMID:26954467

  20. ThERF1 regulates its target genes via binding to a novel cis-acting element in response to salt stress

    Institute of Scientific and Technical Information of China (English)

    Liuqiang Wang; Chao Wang; Liping Qin; Wenjin Liu; Yucheng Wang

    2015-01-01

    Ethylene responsive factors (ERFs) are plant-specific transcription factors that are involved in a variety of biological processes. We previously demonstrated that an ERF gene from Tamarix hispida, ThERF1, encodes a protein binding to GCC-box and DRE motifs and negatively modulates abiotic stress tolerance. In the present study, microarray analysis was performed to study the genes regulated by ThERF1 on a genomic scale. There were 154 and 307 genes (respectively representing 134 and 260 unique genes) significantly up-and downregulated by ThERF1 under salt stress conditions, respectively. A novel motif, named TTG, was identified to be recognized by ThERF1, which commonly presents in the promoters of ThERF1-targeted genes. The TTG motif is also bound by other ERFs of a different subfamily from T. hispida and Arabidopsis, indicating that it is commonly recognized by ERF proteins. The binding affinities of ERFs to the TTG motif are significantly induced by salt stress. The TTG motif is more enriched than the GCC-box and DRE motifs in the promoters of ThERF1-targeted genes. Taken together, these studies suggested that the TTG motif plays an important role in the gene expression regulated by ERFs in response to salt stress.

  1. Affine and degenerate affine BMW algebras: The center

    CERN Document Server

    Daugherty, Zajj; Virk, Rahbar

    2011-01-01

    The degenerate affine and affine BMW algebras arise naturally in the context of Schur-Weyl duality for orthogonal and symplectic Lie algebras and quantum groups, respectively. Cyclotomic BMW algebras, affine Hecke algebras, cyclotomic Hecke algebras, and their degenerate versions are quotients. In this paper the theory is unified by treating the orthogonal and symplectic cases simultaneously; we make an exact parallel between the degenerate affine and affine cases via a new algebra which takes the role of the affine braid group for the degenerate setting. A main result of this paper is an identification of the centers of the affine and degenerate affine BMW algebras in terms of rings of symmetric functions which satisfy a "cancellation property" or "wheel condition" (in the degenerate case, a reformulation of a result of Nazarov). Miraculously, these same rings also arise in Schubert calculus, as the cohomology and K-theory of isotropic Grassmanians and symplectic loop Grassmanians. We also establish new inte...

  2. Targeting amino acid metabolism in cancer growth and anti-tumor immune response

    Institute of Scientific and Technical Information of China (English)

    Elitsa; Ananieva

    2015-01-01

    Recent advances in amino acid metabolism have revealed that targeting amino acid metabolic enzymes in cancer therapy is a promising strategy for the development of novel therapeutic agents. There are currently several drugs in clinical trials that specifically target amino acid metabolic pathways in tumor cells. In the context of the tumor microenvironment,however,tumor cells form metabolic relationships with immune cells,and they oftencompete for common nutrients. Many tumors evolved to escape immune surveillance by taking advantage of their metabolic flexibility and redirecting nutrients for their own advantage. This review outlines the most recent advances in targeting amino acid metabolic pathways in cancer therapy while giving consideration to the impact these pathways may have on the anti-tumor immune response.

  3. Vascular endothelium as a target of immune response in renal transplant rejection.

    Science.gov (United States)

    Piotti, Giovanni; Palmisano, Alessandra; Maggiore, Umberto; Buzio, Carlo

    2014-01-01

    This review of clinical and experimental studies aims at analyzing the interplay between graft endothelium and host immune system in renal transplantation, and how it affects the survival of the graft. Graft endothelium is indeed the first barrier between self and non-self that is encountered by host lymphocytes upon reperfusion of vascularized solid transplants. Endothelial cells (EC) express all the major sets of antigens (Ag) that elicit host immune response, and therefore represent a preferential target in organ rejection. Some of the Ag expressed by EC are target of the antibody-mediated response, such as the AB0 blood group system, the human leukocyte antigens (HLA), and MHC class I related chain A antigens (MICA) systems, and the endothelial cell-restricted Ag; for each of these systems, the mechanisms of interaction and damage of both preformed and de novo donor-specific antibodies are reviewed along with their impact on renal graft survival. Moreover, the rejection process can force injured EC to expose cryptic self-Ag, toward which an autoimmune response mounts, overlapping to the allo-immune response in the damaging of the graft. Not only are EC a passive target of the host immune response but also an active player in lymphocyte activation; therefore, their interaction with allogenic T-cells is analyzed on the basis of experimental in vitro and in vivo studies, according to the patterns of expression of the HLA class I and II and the co-stimulatory molecules specific for cytotoxic and helper T-cells. Finally, as the response that follows transplantation has proven to be not necessarily destructive, the factors that foster graft endothelium functioning in spite of rejection, and how they could be therapeutically harnessed to promote long-term graft acceptance, are described: accommodation that is resistance of EC to donor-specific antibodies, and endothelial cell ability to induce Foxp3+ regulatory T-cells, that are crucial mediators of tolerance. PMID

  4. Hierarchical Affinity Propagation

    CERN Document Server

    Givoni, Inmar; Frey, Brendan J

    2012-01-01

    Affinity propagation is an exemplar-based clustering algorithm that finds a set of data-points that best exemplify the data, and associates each datapoint with one exemplar. We extend affinity propagation in a principled way to solve the hierarchical clustering problem, which arises in a variety of domains including biology, sensor networks and decision making in operational research. We derive an inference algorithm that operates by propagating information up and down the hierarchy, and is efficient despite the high-order potentials required for the graphical model formulation. We demonstrate that our method outperforms greedy techniques that cluster one layer at a time. We show that on an artificial dataset designed to mimic the HIV-strain mutation dynamics, our method outperforms related methods. For real HIV sequences, where the ground truth is not available, we show our method achieves better results, in terms of the underlying objective function, and show the results correspond meaningfully to geographi...

  5. Antibody affinity maturation

    DEFF Research Database (Denmark)

    Skjødt, Mette Louise

    linker for yeast surface display of scFv and scFab fragments, we compared a series of different Gly-Ser-based linkers in display and antigen binding proficiency. We show that these formats of the model antibody can accommodate linkers of different lengths and that introduction of alanine or glutamate......-2. Based on the presented data we suggest that affinity maturation of the model antibody proceeds through multiple incremental steps of subtle improvements. We moreover conclude that the best affinity improved candidates are likely to be obtained through optimization of both the antigen...... fragments by in vivo homologous recombination large combinatorial antibody libraries can easily be generated. We have optimized ordered assembly of three CDR fragments into a gapped vector and observed increased transformation efficiency in a yeast strain carrying a deletion of the SGS1 helicase...

  6. Gaussian Affine Feature Detector

    OpenAIRE

    Xu, Xiaopeng; Zhang, Xiaochun

    2011-01-01

    A new method is proposed to get image features' geometric information. Using Gaussian as an input signal, a theoretical optimal solution to calculate feature's affine shape is proposed. Based on analytic result of a feature model, the method is different from conventional iterative approaches. From the model, feature's parameters such as position, orientation, background luminance, contrast, area and aspect ratio can be extracted. Tested with synthesized and benchmark data, the method achieve...

  7. Affinity driven social networks

    Science.gov (United States)

    Ruyú, B.; Kuperman, M. N.

    2007-04-01

    In this work we present a model for evolving networks, where the driven force is related to the social affinity between individuals of a population. In the model, a set of individuals initially arranged on a regular ordered network and thus linked with their closest neighbors are allowed to rearrange their connections according to a dynamics closely related to that of the stable marriage problem. We show that the behavior of some topological properties of the resulting networks follows a non trivial pattern.

  8. Dopamine responsiveness is regulated by targeted sorting of D2 receptors

    OpenAIRE

    Bartlett, Selena E.; Enquist, Johan; Hopf, Frederic W.; Lee, Josephine H.; Gladher, Fredrik; Kharazia, Viktor; Waldhoer, Maria; Mailliard, William S.; Armstrong, Randall; Bonci, Antonello; Whistler, Jennifer L.

    2005-01-01

    Aberrant dopaminergic signaling is a critical determinant in multiple psychiatric disorders, and in many disease states, dopamine receptor number is altered. Here we identify a molecular mechanism that selectively targets D2 receptors for degradation after their activation by dopamine. The degradative fate of D2 receptors is determined by an interaction with G protein coupled receptor-associated sorting protein (GASP). As a consequence of this GASP interaction, D2 responses in rat brain fail ...

  9. Generation of T Cell Responses Targeting the Reactive Metabolite of Halothane in Mice

    OpenAIRE

    YOU, QIANG; Cheng, Linling; Ju, Cynthia

    2010-01-01

    Immune-mediated adverse drug reactions (IADRs) represent a significant problem in clinical practice and drug development. Studies of the underlying mechanisms of IADRs have been hampered by the lack of animal models. Halothane causes severe allergic hepatitis with clinical features consistent with an IADR. Our ultimate goal is to develop a mouse model of halothane hepatitis. Evidence suggests that adaptive immune responses targeting liver protein adducts of the reactive metabolite (TFA) play ...

  10. A target based approach identifies genomic predictors of breast cancer patient response to chemotherapy

    Directory of Open Access Journals (Sweden)

    Hallett Robin M

    2012-05-01

    Full Text Available Abstract Background The efficacy of chemotherapy regimens in breast cancer patients is variable and unpredictable. Whether individual patients either achieve long-term remission or suffer recurrence after therapy may be dictated by intrinsic properties of their breast tumors including genetic lesions and consequent aberrant transcriptional programs. Global gene expression profiling provides a powerful tool to identify such tumor-intrinsic transcriptional programs, whose analyses provide insight into the underlying biology of individual patient tumors. For example, multi-gene expression signatures have been identified that can predict the likelihood of disease reccurrence, and thus guide patient prognosis. Whereas such prognostic signatures are being introduced in the clinical setting, similar signatures that predict sensitivity or resistance to chemotherapy are not currently clinically available. Methods We used gene expression profiling to identify genes that were co-expressed with genes whose transcripts encode the protein targets of commonly used chemotherapeutic agents. Results Here, we present target based expression indices that predict breast tumor response to anthracycline and taxane based chemotherapy. Indeed, these signatures were independently predictive of chemotherapy response after adjusting for standard clinic-pathological variables such as age, grade, and estrogen receptor status in a cohort of 488 breast cancer patients treated with adriamycin and taxotere/taxol. Conclusions Importantly, our findings suggest the practicality of developing target based indices that predict response to therapeutics, as well as highlight the possibility of using gene signatures to guide the use of chemotherapy during treatment of breast cancer patients.

  11. Mitochondrial targeting of bilirubin regulatory enzymes: An adaptive response to oxidative stress

    Energy Technology Data Exchange (ETDEWEB)

    Muhsain, Siti Nur Fadzilah, E-mail: sitinurfadzilah077@ppinang.uitm.edu.my [The University of Queensland, National Research Centre for Environmental Toxicology (Entox), 4072 Brisbane, Queensland (Australia); Faculty of Pharmacy, University Teknologi Mara (Malaysia); Lang, Matti A., E-mail: m.lang@uq.edu.au [The University of Queensland, National Research Centre for Environmental Toxicology (Entox), 4072 Brisbane, Queensland (Australia); Abu-Bakar, A' edah, E-mail: a.abubakar@uq.edu.au [The University of Queensland, National Research Centre for Environmental Toxicology (Entox), 4072 Brisbane, Queensland (Australia)

    2015-01-01

    The intracellular level of bilirubin (BR), an endogenous antioxidant that is cytotoxic at high concentrations, is tightly controlled within the optimal therapeutic range. We have recently described a concerted intracellular BR regulation by two microsomal enzymes: heme oxygenase 1 (HMOX1), essential for BR production and cytochrome P450 2A5 (CYP2A5), a BR oxidase. Herein, we describe targeting of these enzymes to hepatic mitochondria during oxidative stress. The kinetics of microsomal and mitochondrial BR oxidation were compared. Treatment of DBA/2J mice with 200 mg pyrazole/kg/day for 3 days increased hepatic intracellular protein carbonyl content and induced nucleo-translocation of Nrf2. HMOX1 and CYP2A5 proteins and activities were elevated in microsomes and mitoplasts but not the UGT1A1, a catalyst of BR glucuronidation. A CYP2A5 antibody inhibited 75% of microsomal BR oxidation. The inhibition was absent in control mitoplasts but elevated to 50% after treatment. An adrenodoxin reductase antibody did not inhibit microsomal BR oxidation but inhibited 50% of mitochondrial BR oxidation. Ascorbic acid inhibited 5% and 22% of the reaction in control and treated microsomes, respectively. In control mitoplasts the inhibition was 100%, which was reduced to 50% after treatment. Bilirubin affinity to mitochondrial and microsomal CYP2A5 enzyme is equally high. Lastly, the treatment neither released cytochrome c into cytoplasm nor dissipated membrane potential, indicating the absence of mitochondrial membrane damage. Collectively, the observations suggest that BR regulatory enzymes are recruited to mitochondria during oxidative stress and BR oxidation by mitochondrial CYP2A5 is supported by mitochondrial mono-oxygenase system. The induced recruitment potentially confers membrane protection. - Highlights: • Pyrazole induces oxidative stress in the mouse liver. • Pyrazole-induced oxidative stress induces mitochondrial targeting of key bilirubin regulatory enzymes, HMOX1

  12. European and international collaboration in affinity proteomics.

    Science.gov (United States)

    Stoevesandt, Oda; Taussig, Michael J

    2012-06-15

    In affinity proteomics, specific protein-binding molecules (a.k.a. binders), principally antibodies, are applied as reagents in proteome analysis. In recent years, advances in binder technologies have created the potential for an unprecedented view on protein expression and distribution patterns in plasma, cells and tissues and increasingly on protein function. Particular strengths of affinity proteomics methods include detecting proteins in their natural environments of cell or tissue, high sensitivity and selectivity for detection of low abundance proteins and exploiting binding actions such as functional interference in living cells. To maximise the use and impact of affinity reagents, it will be essential to create comprehensive, standardised binder collections. With this in mind, the EU FP7 programme AFFINOMICS (http://www.affinomics.org), together with the preceding EU programmes ProteomeBinders and AffinityProteome, aims to extend affinity proteomics research by generating a large-scale resource of validated protein-binding molecules for characterisation of the human proteome. Activity is directed at producing binders to about 1000 protein targets, primarily in signal transduction and cancer, by establishing a high throughput, coordinated production pipeline. An important aspect of AFFINOMICS is the development of highly efficient recombinant selection methods, based on phage, cell and ribosome display, capable of producing high quality binders at greater throughput and lower cost than hitherto. The programme also involves development of innovative and sensitive technologies for specific detection of target proteins and their interactions, and deployment of binders in proteomics studies of clinical relevance. The need for such binder generation programmes is now recognised internationally, with parallel initiatives in the USA for cancer (NCI) and transcription factors (NIH) and within the Human Proteome Organisation (HUPO). The papers in this volume of New

  13. Surface Modified Multifunctional and Stimuli Responsive Nanoparticles for Drug Targeting: Current Status and Uses.

    Science.gov (United States)

    Siafaka, Panoraia I; Üstündağ Okur, Neslihan; Karavas, Evangelos; Bikiaris, Dimitrios N

    2016-01-01

    Nanocarriers, due to their unique features, are of increased interest among researchers working with pharmaceutical formulations. Polymeric nanoparticles and nanocapsules, involving non-toxic biodegradable polymers, liposomes, solid lipid nanoparticles, and inorganic-organic nanomaterials, are among the most used carriers for drugs for a broad spectrum of targeted diseases. In fact, oral, injectable, transdermal-dermal and ocular formulations mainly consist of the aforementioned nanomaterials demonstrating promising characteristics such as long circulation, specific targeting, high drug loading capacity, enhanced intracellular penetration, and so on. Over the last decade, huge advances in the development of novel, safer and less toxic nanocarriers with amended properties have been made. In addition, multifunctional nanocarriers combining chemical substances, vitamins and peptides via coupling chemistry, inorganic particles coated by biocompatible materials seem to play a key role considering that functionalization can enhance characteristics such as biocompatibility, targetability, environmental friendliness, and intracellular penetration while also have limited side effects. This review aims to summarize the "state of the art" of drug delivery carriers in nanosize, paying attention to their surface functionalization with ligands and other small or polymeric compounds so as to upgrade active and passive targeting, different release patterns as well as cell targeting and stimuli responsibility. Lastly, future aspects and potential uses of nanoparticulated drug systems are outlined. PMID:27589733

  14. The CERN antiproton target: hydrocode analysis of its core material dynamic response under proton beam impact

    CERN Document Server

    Martin, Claudio Torregrosa; Calviani, Marco; Muñoz-Cobo, José-Luis

    2016-01-01

    Antiprotons are produced at CERN by colliding a 26 GeV/c proton beam with a fixed target made of a 3 mm diameter, 55 mm length iridium core. The inherent characteristics of antiproton production involve extremely high energy depositions inside the target when impacted by each primary proton beam, making it one of the most dynamically demanding among high energy solid targets in the world, with a rise temperature above 2000 {\\deg}C after each pulse impact and successive dynamic pressure waves of the order of GPa's. An optimized redesign of the current target is foreseen for the next 20 years of operation. As a first step in the design procedure, this numerical study delves into the fundamental phenomena present in the target material core under proton pulse impact and subsequent pressure wave propagation by the use of hydrocodes. Three major phenomena have been identified, (i) the dominance of a high frequency radial wave which produces destructive compressive-to-tensile pressure response (ii) The existence of...

  15. Affine morphisms at zero level

    CERN Document Server

    Das, Paramita; Gupta, Ved Prakash

    2010-01-01

    Given a finite index subfactor, we show that the {\\em affine morphisms at zero level} in the affine category over the planar algebra associated to the subfactor is isomorphic to the fusion algebra of the subfactor as a *-algebra.

  16. On the Affine Isoperimetric Inequalities

    Indian Academy of Sciences (India)

    Wuyang Yu; Gangsong Leng

    2011-11-01

    We obtain an isoperimetric inequality which estimate the affine invariant -surface area measure on convex bodies. We also establish the reverse version of -Petty projection inequality and an affine isoperimetric inequality of $_{-p}K$.

  17. Circular revisit orbits design for responsive mission over a single target

    Science.gov (United States)

    Li, Taibo; Xiang, Junhua; Wang, Zhaokui; Zhang, Yulin

    2016-10-01

    The responsive orbits play a key role in addressing the mission of Operationally Responsive Space (ORS) because of their capabilities. These capabilities are usually focused on supporting specific targets as opposed to providing global coverage. One subtype of responsive orbits is repeat coverage orbit which is nearly circular in most remote sensing applications. This paper deals with a special kind of repeating ground track orbit, referred to as circular revisit orbit. Different from traditional repeat coverage orbits, a satellite on circular revisit orbit can visit a target site at both the ascending and descending stages in one revisit cycle. This typology of trajectory allows a halving of the traditional revisit time and does a favor to get useful information for responsive applications. However the previous reported numerical methods in some references often cost lots of computation or fail to obtain such orbits. To overcome this difficulty, an analytical method to determine the existence conditions of the solutions to revisit orbits is presented in this paper. To this end, the mathematical model of circular revisit orbit is established under the central gravity model and the J2 perturbation. A constraint function of the circular revisit orbit is introduced, and the monotonicity of that function has been studied. The existent conditions and the number of such orbits are naturally worked out. Taking the launch cost into consideration, optimal design model of circular revisit orbit is established to achieve a best orbit which visits a target twice a day in the morning and in the afternoon respectively for several days. The result shows that it is effective to apply circular revisit orbits in responsive application such as reconnoiter of natural disaster.

  18. Adjoint affine fusion and tadpoles

    OpenAIRE

    Urichuk, Andrew; Walton, Mark A.

    2016-01-01

    We study affine fusion with the adjoint representation. For simple Lie algebras, elementary and universal formulas determine the decomposition of a tensor product of an integrable highest-weight representation with the adjoint representation. Using the (refined) affine depth rule, we prove that equally striking results apply to adjoint affine fusion. For diagonal fusion, a coefficient equals the number of nonzero Dynkin labels of the relevant affine highest weight, minus 1. A nice lattice-pol...

  19. Affine Patches on Positroid Varieties and Affine Pipe Dreams (Thesis)

    CERN Document Server

    Snider, Michelle

    2010-01-01

    The objects of interest in this thesis are positroid varieties in the Grassmannian, which are indexed by juggling patterns. In particular, we study affine patches on these positroid varieties. Our main result corresponds these affine patches to Kazhdan-Lusztig varieties in the affine Grassmannian. We develop a new term order and study how these spaces are related to subword complexes and Stanley-Reisner ideals. We define an extension of pipe dreams to the affine case and conclude by showing how our affine pipe dreams are generalizations of Cauchon and Le diagrams.

  20. Affine and quasi-affine frames for rational dilations

    DEFF Research Database (Denmark)

    Bownik, Marcin; Lemvig, Jakob

    2011-01-01

    , the corresponding family of quasi-affine systems are frames with uniform frame bounds. We also prove a similar equivalence result between pairs of dual affine frames and dual quasi-affine frames. Finally, we uncover some fundamental differences between the integer and rational settings by exhibiting an example......In this paper we extend the investigation of quasi-affine systems, which were originally introduced by Ron and Shen [J. Funct. Anal. 148 (1997), 408-447] for integer, expansive dilations, to the class of rational, expansive dilations. We show that an affine system is a frame if, and only if...

  1. Inhibition of MNK pathways enhances cancer cell response to chemotherapy with temozolomide and targeted radionuclide therapy.

    Science.gov (United States)

    Grzmil, Michal; Seebacher, Jan; Hess, Daniel; Behe, Martin; Schibli, Roger; Moncayo, Gerald; Frank, Stephan; Hemmings, Brian A

    2016-09-01

    Current standard-of-care treatment for malignant cancers includes radiotherapy and adjuvant chemotherapy. Here, we report increased MAP kinase-interacting kinase (MNK)-regulated phosphorylation of translation initiation factor 4E (eIF4E) in glioma cells upon temozolomide (TMZ) treatment and in medullary thyroid carcinoma (MTC) cells in response to targeted radionuclide therapy. Depletion of MNK activity by using two MNK inhibitors, CGP57380 or cercosporamide, as well as by MNK1-specific knockdown sensitized glioblastoma (GBM) cells and GBM-derived spheres to TMZ. Furthermore, CGP57380 treatment enhanced response of MTC cells to (177)Lu-labeled gastrin analogue. In order to understand how MNK signaling pathways support glioma survival we analyzed putative MNK substrates by quantitative phosphoproteomics in normal condition and in the presence of TMZ. We identified MNK inhibitor-sensitive phosphorylation sites on eIF4G1, mutations of which either influenced eIF4E phosphorylation or glioma cell response to TMZ, pointing to altered regulation of translation initiation as a resistance mechanism. Pharmacological inhibition of overexpressed MNK1 by CGP57380 reduced eIF4E phosphorylation and induced association of inactive MNK1 with eIF4G1. Taken together, our data show an activation of MNK-mediated survival mechanisms in response to either glioma chemotherapy or MTC targeted radiation and suggest that inhibition of MNK activity represents an attractive sensitizing strategy for cancer treatments.

  2. Inhibition of MNK pathways enhances cancer cell response to chemotherapy with temozolomide and targeted radionuclide therapy.

    Science.gov (United States)

    Grzmil, Michal; Seebacher, Jan; Hess, Daniel; Behe, Martin; Schibli, Roger; Moncayo, Gerald; Frank, Stephan; Hemmings, Brian A

    2016-09-01

    Current standard-of-care treatment for malignant cancers includes radiotherapy and adjuvant chemotherapy. Here, we report increased MAP kinase-interacting kinase (MNK)-regulated phosphorylation of translation initiation factor 4E (eIF4E) in glioma cells upon temozolomide (TMZ) treatment and in medullary thyroid carcinoma (MTC) cells in response to targeted radionuclide therapy. Depletion of MNK activity by using two MNK inhibitors, CGP57380 or cercosporamide, as well as by MNK1-specific knockdown sensitized glioblastoma (GBM) cells and GBM-derived spheres to TMZ. Furthermore, CGP57380 treatment enhanced response of MTC cells to (177)Lu-labeled gastrin analogue. In order to understand how MNK signaling pathways support glioma survival we analyzed putative MNK substrates by quantitative phosphoproteomics in normal condition and in the presence of TMZ. We identified MNK inhibitor-sensitive phosphorylation sites on eIF4G1, mutations of which either influenced eIF4E phosphorylation or glioma cell response to TMZ, pointing to altered regulation of translation initiation as a resistance mechanism. Pharmacological inhibition of overexpressed MNK1 by CGP57380 reduced eIF4E phosphorylation and induced association of inactive MNK1 with eIF4G1. Taken together, our data show an activation of MNK-mediated survival mechanisms in response to either glioma chemotherapy or MTC targeted radiation and suggest that inhibition of MNK activity represents an attractive sensitizing strategy for cancer treatments. PMID:27289018

  3. The effects of incidentally learned temporal and spatial predictability on response times and visual fixations during target detection and discrimination.

    Science.gov (United States)

    Beck, Melissa R; Hong, S Lee; van Lamsweerde, Amanda E; Ericson, Justin M

    2014-01-01

    Responses are quicker to predictable stimuli than if the time and place of appearance is uncertain. Studies that manipulate target predictability often involve overt cues to speed up response times. However, less is known about whether individuals will exhibit faster response times when target predictability is embedded within the inter-trial relationships. The current research examined the combined effects of spatial and temporal target predictability on reaction time (RT) and allocation of overt attention in a sustained attention task. Participants responded as quickly as possible to stimuli while their RT and eye movements were measured. Target temporal and spatial predictability were manipulated by altering the number of: 1) different time intervals between a response and the next target; and 2) possible spatial locations of the target. The effects of target predictability on target detection (Experiment 1) and target discrimination (Experiment 2) were tested. For both experiments, shorter RTs as target predictability increased across both space and time were found. In addition, the influences of spatial and temporal target predictability on RT and the overt allocation of attention were task dependent; suggesting that effective orienting of attention relies on both spatial and temporal predictability. These results indicate that stimulus predictability can be increased without overt cues and detected purely through inter-trial relationships over the course of repeated stimulus presentations.

  4. The effects of incidentally learned temporal and spatial predictability on response times and visual fixations during target detection and discrimination.

    Directory of Open Access Journals (Sweden)

    Melissa R Beck

    Full Text Available Responses are quicker to predictable stimuli than if the time and place of appearance is uncertain. Studies that manipulate target predictability often involve overt cues to speed up response times. However, less is known about whether individuals will exhibit faster response times when target predictability is embedded within the inter-trial relationships. The current research examined the combined effects of spatial and temporal target predictability on reaction time (RT and allocation of overt attention in a sustained attention task. Participants responded as quickly as possible to stimuli while their RT and eye movements were measured. Target temporal and spatial predictability were manipulated by altering the number of: 1 different time intervals between a response and the next target; and 2 possible spatial locations of the target. The effects of target predictability on target detection (Experiment 1 and target discrimination (Experiment 2 were tested. For both experiments, shorter RTs as target predictability increased across both space and time were found. In addition, the influences of spatial and temporal target predictability on RT and the overt allocation of attention were task dependent; suggesting that effective orienting of attention relies on both spatial and temporal predictability. These results indicate that stimulus predictability can be increased without overt cues and detected purely through inter-trial relationships over the course of repeated stimulus presentations.

  5. Nanoparticles responsive to the inflammatory microenvironment for targeted treatment of arterial restenosis.

    Science.gov (United States)

    Feng, Shibin; Hu, Ying; Peng, Song; Han, Songling; Tao, Hui; Zhang, Qixiong; Xu, Xiaoqiu; Zhang, Jianxiang; Hu, Houyuan

    2016-10-01

    Coronary arterial disease (CAD) remains the leading cause of death globally. Percutaneous coronary interventions are frequently used nonsurgical techniques for treating CAD, which may unfortunately lead to arterial restenosis. Currently, there are no effective drugs that can thoroughly prevent restenosis. We hypothesize inflammation-triggerable nanomedicines may function as effective therapeutics for targeted therapy of restenosis, by preferentially releasing their payload at the diseased site. To demonstrate our hypothesis and develop targeted nanotherapies for restenosis, this study was designed to examine effectiveness of nanomedicines responsive to the inflammatory microenvironment with mild acidity and high reactive oxygen species (ROS). To this end, an acetalated β-cyclodextrin (β-CD) material (Ac-bCD) was synthesized as a pH-responsive carrier material, while a ROS-responsive material (Ox-bCD) was produced by hydrophobic functionalization of β-CD with an oxidation-labile group. Based on these two responsive materials, either pH- or ROS-responsive nanoparticles (NPs) were produced by a nanoprecipitation technique and fully characterized. Using rapamycin (RAP) as a candidate drug, responsive nanotherapies were fabricated. In vitro hydrolysis and release studies confirmed these nanovehicles and nanotherapies exhibited desirable responsive behaviors. Both in vitro cell culture and in vivo evaluations revealed their good safety profile. These responsive NPs could be effectively internalized by rat vascular smooth muscle cells, which in turn notably potentiated anti-proliferation and anti-migration activities of RAP. After intravenous (i.v.) injection, NPs may be accumulated at the injured site in the carotid artery of rats subjected to balloon angioplasty injury. Compared with a non-responsive nanotherapy based on poly(lactide-co-glycolide), treatment with either pH- or ROS-responsive nanotherapy by i.v. injection more effectively attenuated neointimal

  6. MPLA incorporation into DC-targeting glycoliposomes favours anti-tumour T cell responses.

    Science.gov (United States)

    Boks, Martine A; Ambrosini, Martino; Bruijns, Sven C; Kalay, Hakan; van Bloois, Louis; Storm, Gert; Garcia-Vallejo, Juan J; van Kooyk, Yvette

    2015-10-28

    Dendritic cells (DC) are attractive targets for cancer immunotherapy as they initiate strong and long-lived tumour-specific T cell responses. DC can be effectively targeted in vivo with tumour antigens by using nanocarriers such as liposomes. Cross-presentation of tumour antigens is enhanced with strong adjuvants such as TLR ligands. However, often these adjuvants have off-target effects, and would benefit from a DC-specific targeting strategy, similar to the tumour antigen. The goal of this study was to develop a strategy for specifically targeting DC with tumour antigen and adjuvant by using glycoliposomes. We have generated liposomes containing the glycan Lewis(Le)(X) which is highly specific for the C-type lectin receptor DC-SIGN expressed by DC. Le(X)-modified liposomes were taken up by human monocyte-derived DC in a DC-SIGN-specific manner. As adjuvants we incorporated the TLR ligands Pam3CySK4, Poly I:C, MPLA and R848 into liposomes and compared their adjuvant capacity on DC. Incorporation of the TLR4 ligand MPLA into glycoliposomes induced DC maturation and production of pro-inflammatory cytokines, in a DC-SIGN-specific manner, and DC activation was comparable to administration of soluble MPLA. Incorporation of MPLA into glycoliposomes significantly enhanced antigen cross-presentation of the melanoma tumour antigen gp100280-288 peptide to CD8(+) T cells compared to non-glycosylated MPLA liposomes. Importantly, antigen cross-presentation of the gp100280-288 peptide was significantly higher using MPLA glycoliposomes compared to the co-administration of soluble MPLA with glycoliposomes. Taken together, our data demonstrates that specific targeting of a gp100 tumour antigen and the adjuvant MPLA to DC-SIGN-expressing DC enhances the uptake of peptide-containing liposomes, the activation of DC, and induces tumour antigen-specific CD8(+) T cell responses. These data demonstrate that adjuvant-containing glycoliposome-based vaccines targeting DC-SIGN(+) DC

  7. Assessing the role of alternative response rates and reinforcer rates in resistance to extinction of target responding when combining stimuli.

    Science.gov (United States)

    Podlesnik, Christopher A; Bai, John Y H; Skinner, Katherine A

    2016-05-01

    Studies of behavioral momentum reveal that reinforcing an alternative response in the presence of a target response reduces the rate of target responding but increases its persistence, relative to training the target response on its own. Because of the parallels between these studies and differential-reinforcement techniques to reduce problem behavior in clinical settings, alternative techniques to reduce problem behavior without enhancing its persistence are being explored. One potential solution is to train an alternative response in a separate stimulus context from problem behavior before combining the alternative stimulus with the target stimulus. The present study assessed how differences in reinforcement contingencies and rate for alternative responding influenced resistance to extinction of target responding when combining alternative and target stimuli in pigeons. Across three experiments, alternative stimuli signaling a response-reinforcer dependency and greater reinforcer rates more effectively decreased the persistence of target responding when combining alternative and target stimuli within the same extinction tests, but not when compared across separate extinction tests. Overall, these findings reveal that differences in competition between alternative and target responding produced by contingencies of alternative reinforcement could influence the effectiveness of treating problem behavior through combining stimulus contexts. PMID:27193243

  8. Cellular Pathways in Response to Ionizing Radiation and Their Targetability for Tumor Radiosensitization.

    Science.gov (United States)

    Maier, Patrick; Hartmann, Linda; Wenz, Frederik; Herskind, Carsten

    2016-01-14

    During the last few decades, improvements in the planning and application of radiotherapy in combination with surgery and chemotherapy resulted in increased survival rates of tumor patients. However, the success of radiotherapy is impaired by two reasons: firstly, the radioresistance of tumor cells and, secondly, the radiation-induced damage of normal tissue cells located in the field of ionizing radiation. These limitations demand the development of drugs for either radiosensitization of tumor cells or radioprotection of normal tissue cells. In order to identify potential targets, a detailed understanding of the cellular pathways involved in radiation response is an absolute requirement. This review describes the most important pathways of radioresponse and several key target proteins for radiosensitization.

  9. Cellular Pathways in Response to Ionizing Radiation and Their Targetability for Tumor Radiosensitization

    Directory of Open Access Journals (Sweden)

    Patrick Maier

    2016-01-01

    Full Text Available During the last few decades, improvements in the planning and application of radiotherapy in combination with surgery and chemotherapy resulted in increased survival rates of tumor patients. However, the success of radiotherapy is impaired by two reasons: firstly, the radioresistance of tumor cells and, secondly, the radiation-induced damage of normal tissue cells located in the field of ionizing radiation. These limitations demand the development of drugs for either radiosensitization of tumor cells or radioprotection of normal tissue cells. In order to identify potential targets, a detailed understanding of the cellular pathways involved in radiation response is an absolute requirement. This review describes the most important pathways of radioresponse and several key target proteins for radiosensitization.

  10. In situ crosslinked smart polypeptide nanoparticles for multistage responsive tumor-targeted drug delivery

    Science.gov (United States)

    Yi, Huqiang; Liu, Peng; Sheng, Nan; Gong, Ping; Ma, Yifan; Cai, Lintao

    2016-03-01

    Smart tumor-targeted drug delivery is crucial for improving the effect of chemotherapy and reducing the adverse effects. Here, we synthesized a smart polypeptide copolymer based on n-butylamine-poly(l-lysine)-b-poly(l-cysteine) (PLL-PLC) with functionalization of folic acid (FA) and 1,2-dicarboxylic-cyclohexene anhydride (DCA) for multistage responsive tumor-targeted drug delivery. The copolymers (FA-PLL(DCA)-PLC) spontaneously crosslinked in situ to form redox and pH dual responsive FA-PLL(DCA)-PLC nanoparticles (FD-NPs), which had a reversible zeta potential around -30 mV at pH 7.4, but switched to +15 mV at pH 5.0. Moreover, FD-NPs effectively loaded DOX with a loading capacity at 15.7 wt%. At pH 7.4, only 24.5% DOX was released within 60 h. However, at pH 5.0, the presence of 10 mM DTT dramatically accelerated DOX release with over 90% of DOX released within 10 h. Although the FD-NPs only enhanced DOX uptake in FA receptor positive (FR+) cancer cells at pH 7.4, a weak acidic condition promoted FD-NP-facilitated DOX uptake in both FR+ HeLa and FR- A549 cells, as well as significantly improving cellular binding and end/lysosomal escape. In vivo studies in a HeLa cancer model demonstrated that the charge-reversible FD-NPs delivered DOX into tumors more effectively than charge-irreversible nanoparticles. Hence, these multistage responsive FD-NPs would serve as highly efficient drug vectors for targeted cancer chemotherapy.Smart tumor-targeted drug delivery is crucial for improving the effect of chemotherapy and reducing the adverse effects. Here, we synthesized a smart polypeptide copolymer based on n-butylamine-poly(l-lysine)-b-poly(l-cysteine) (PLL-PLC) with functionalization of folic acid (FA) and 1,2-dicarboxylic-cyclohexene anhydride (DCA) for multistage responsive tumor-targeted drug delivery. The copolymers (FA-PLL(DCA)-PLC) spontaneously crosslinked in situ to form redox and pH dual responsive FA-PLL(DCA)-PLC nanoparticles (FD-NPs), which had a reversible

  11. Broad-spectrum anti-biofilm peptide that targets a cellular stress response.

    Directory of Open Access Journals (Sweden)

    César de la Fuente-Núñez

    2014-05-01

    Full Text Available Bacteria form multicellular communities known as biofilms that cause two thirds of all infections and demonstrate a 10 to 1000 fold increase in adaptive resistance to conventional antibiotics. Currently, there are no approved drugs that specifically target bacterial biofilms. Here we identified a potent anti-biofilm peptide 1018 that worked by blocking (pppGpp, an important signal in biofilm development. At concentrations that did not affect planktonic growth, peptide treatment completely prevented biofilm formation and led to the eradication of mature biofilms in representative strains of both Gram-negative and Gram-positive bacterial pathogens including Pseudomonas aeruginosa, Escherichia coli, Acinetobacter baumannii, Klebsiella pneumoniae, methicillin resistant Staphylococcus aureus, Salmonella Typhimurium and Burkholderia cenocepacia. Low levels of the peptide led to biofilm dispersal, while higher doses triggered biofilm cell death. We hypothesized that the peptide acted to inhibit a common stress response in target species, and that the stringent response, mediating (pppGpp synthesis through the enzymes RelA and SpoT, was targeted. Consistent with this, increasing (pppGpp synthesis by addition of serine hydroxamate or over-expression of relA led to reduced susceptibility to the peptide. Furthermore, relA and spoT mutations blocking production of (pppGpp replicated the effects of the peptide, leading to a reduction of biofilm formation in the four tested target species. Also, eliminating (pppGpp expression after two days of biofilm growth by removal of arabinose from a strain expressing relA behind an arabinose-inducible promoter, reciprocated the effect of peptide added at the same time, leading to loss of biofilm. NMR and chromatography studies showed that the peptide acted on cells to cause degradation of (pppGpp within 30 minutes, and in vitro directly interacted with ppGpp. We thus propose that 1018 targets (pppGpp and marks it for

  12. Synthesis of tetracycline analogs and their bone affinities

    Institute of Scientific and Technical Information of China (English)

    Wen Cai Huang; Hu Zheng; Ling Ling Weng

    2008-01-01

    Tetracycline analogs were designed and synthesized and their bone affinities were tested on hydroxyapatite. The results showedthat the carbonyl-amide-enol structure in A ring and phenol-ketone structure in BCD ring may be responsible for tetracycline's highbone affinity and either A ring or BCD ring has a planar conformation is essential.

  13. Design of Student Information Management Database Application System for Office and Departmental Target Responsibility System

    Science.gov (United States)

    Zhou, Hui

    It is the inevitable outcome of higher education reform to carry out office and departmental target responsibility system, in which statistical processing of student's information is an important part of student's performance review. On the basis of the analysis of the student's evaluation, the student information management database application system is designed by using relational database management system software in this paper. In order to implement the function of student information management, the functional requirement, overall structure, data sheets and fields, data sheet Association and software codes are designed in details.

  14. Watch the Target! Effects in the Affective Misattribution Procedure Become Weaker (But Not Eliminated When Participants Are Motivated to Provide Accurate Responses to the Target

    Directory of Open Access Journals (Sweden)

    Andreas B Eder

    2015-09-01

    Full Text Available Previous research showed that priming effects in the affective misattribution procedure (AMP are unaffected by direct warnings to avoid an influence of the primes. The present research examined whether a priming influence is diminished by task procedures that encourage accurate judgments of the targets. Participants were motivated to categorize the affective meaning of nonsense targets accurately by being made to believe that a true word was presented in each trial and by providing feedback on (allegedly incorrect responses. This condition produced robust priming effects. Priming was however reduced and less reliable relative to more typical AMP conditions in which participants guessed the meaning of openly presented nonsense targets. Affective judgments of nonsense targets were not affected by advance knowledge of the response mapping during the priming phase, which argues against a response-priming explanation of AMP effects. These findings show that affective primes influence evaluative judgments even in conditions in which the motivation to provide accurate responses is high and a priming of motor responses is not possible. Priming effects were however weaker with high accuracy motivation, suggesting that a focus on accurate judgments is an effective strategy to control for an unwanted priming influence in the AMP.

  15. Affinity Purification of Insulin by Peptide-Ligand Affinity Chromatography

    Institute of Scientific and Technical Information of China (English)

    2007-01-01

    The affinity heptapeptide (HWWWPAS) for insulin, selected from phage display library,was coupled to EAH Sepharose 4B gel and packed to a 1-mL column. The column was used for the affinity purification of insulin from protein mixture and commercial insulin preparation. It was observed that the minor impurity in the commercial insulin was removed by the affinity chromatography. Nearly 40 mg of insulin could be purified with the 1-mL affinity column. The results revealed the high specificity and capacity of the affinity column for insulin purification. Moreover, based on the analysis of the amino acids in the peptide sequence, shorter peptides were designed and synthesized for insulin chromatography. As a result, HWWPS was found to be a good alternative to HWWWPAS, while the other two peptides with three or four amino acids showed weak affinity for insulin. The results indicated that the peptide sequence of HWWWPAS was quite conservative for specific binding of insulin.

  16. The Utility of Affine Variables and Affine Coherent States

    CERN Document Server

    Klauder, John R

    2011-01-01

    Affine coherent states are generated by affine kinematical variables much like canonical coherent states are generated by canonical kinematical variables. Although all classical and quantum formalisms normally entail canonical variables, it is shown that affine variables can serve equally well for many classical and quantum studies. This general purpose analysis provides tools to discuss two major applications: (1) the completely successful quantization of a nonrenormalizable scalar quantum field theory by affine techniques, in complete contrast to canonical techniques which only offer triviality; and (2) a formulation of the kinematical portion of quantum gravity that favors affine kinematical variables over canonical kinematical variables, and which generates a framework in which a favorable analysis of the constrained dynamical issues can take place. All this is possible because of the close connection between the affine and the canonical stories, while the few distinctions can be used to advantage when ap...

  17. Ethanol-Induced ADH Activity in Zebrafish: Differential Concentration-Dependent Effects on High- Versus Low-Affinity ADH Enzymes.

    Science.gov (United States)

    Tran, Steven; Nowicki, Magda; Facciol, Amanda; Chatterjee, Diptendu; Gerlai, Robert

    2016-04-01

    Zebrafish express enzymes that metabolize ethanol in a manner comparable to that of mammals, including humans. We previously demonstrated that acute ethanol exposure increases alcohol dehydrogenase (ADH) activity in an inverted U-shaped dose-dependent manner. It was hypothesized that the biphasic dose-response was due to the increased activity of a high-affinity ADH isoform following exposure to low concentrations of ethanol and increased activity of a low-affinity ADH isoform following exposure to higher concentrations of ethanol. To test this hypothesis, we exposed zebrafish to different concentrations of ethanol (0%, 0.25%, 0.5%, and 1.0% v/v) for 30 min and measured the total ADH activity in the zebrafish liver. However, we also repeated this enzyme activity assay using a low concentration of the substrate (ethanol) to determine the activity of high-affinity ADH isoforms. We found that total ADH activity in response to ethanol induces an inverted U-shaped dose-response similar to our previous study. Using a lower substrate level in our enzyme assay targeting high-affinity isozymes, we found a similar dose-response. However, the difference in activity between the high and low substrate assays (high substrate activity - low substrate activity), which provide an index of activity for low-affinity ADH isoforms, revealed no significant effect of ethanol exposure. Our results suggest that the inverted U-shaped dose-response for total ADH activity in response to ethanol is driven primarily by high-affinity isoforms of ADH.

  18. Jacobi Structures on Affine Bundles

    Institute of Scientific and Technical Information of China (English)

    J. GRABOWSKI; D. IGLESIAS; J. C. MARRERO; E. PADR(O)N; P. URBA(N)SKI

    2007-01-01

    We study affine Jacobi structures (brackets) on an affine bundle π: A→M, i.e. Jacobi brackets that close on affine functions. We prove that if the rank of A is non-zero, there is a one-to- one correspondence between affine Jacobi structures on A and Lie algebroid structures on the vector bundle A+=∪p∈M Aff(Ap, R) of affine functionals. In the case rank A = 0, it is shown that there is a one-to-one correspondence between affins Jacobi structures on A and local Lie algebras on A+. Some examples and applications, also for the linear case, are discussed. For a special type of affine Jacobi structures which are canonically exhibited (strongly-affine or affine-homogeneous Jacobi structures) over a real vector space of finite dimension, we describe the leaves of its characteristic foliation as the orbits of an affine representation. These afline Jacobi structures can be viewed as an analog of the Kostant-Arnold-LiouviUe linear Poisson structure on the dual space of a real finite-dimensional Lie algebra.

  19. Genome-wide dynamics of a bacterial response to antibiotics that target the cell envelope

    Directory of Open Access Journals (Sweden)

    Tran Ngat

    2011-05-01

    Full Text Available Abstract Background A decline in the discovery of new antibacterial drugs, coupled with a persistent rise in the occurrence of drug-resistant bacteria, has highlighted antibiotics as a diminishing resource. The future development of new drugs with novel antibacterial activities requires a detailed understanding of adaptive responses to existing compounds. This study uses Streptomyces coelicolor A3(2 as a model system to determine the genome-wide transcriptional response following exposure to three antibiotics (vancomycin, moenomycin A and bacitracin that target distinct stages of cell wall biosynthesis. Results A generalised response to all three antibiotics was identified which involves activation of transcription of the cell envelope stress sigma factor σE, together with elements of the stringent response, and of the heat, osmotic and oxidative stress regulons. Attenuation of this system by deletion of genes encoding the osmotic stress sigma factor σB or the ppGpp synthetase RelA reduced resistance to both vancomycin and bacitracin. Many antibiotic-specific transcriptional changes were identified, representing cellular processes potentially important for tolerance to each antibiotic. Sensitivity studies using mutants constructed on the basis of the transcriptome profiling confirmed a role for several such genes in antibiotic resistance, validating the usefulness of the approach. Conclusions Antibiotic inhibition of bacterial cell wall biosynthesis induces both common and compound-specific transcriptional responses. Both can be exploited to increase antibiotic susceptibility. Regulatory networks known to govern responses to environmental and nutritional stresses are also at the core of the common antibiotic response, and likely help cells survive until any specific resistance mechanisms are fully functional.

  20. Viral Response to Specifically Targeted Antiviral Therapy for Hepatitis C and the Implications for Treatment Success

    Directory of Open Access Journals (Sweden)

    Curtis L Cooper

    2010-01-01

    Full Text Available Currently, hepatitis C virus (HCV antiviral therapy is characterized by long duration, a multitude of side effects, difficult administration and suboptimal success; clearly, alternatives are needed. Collectively, specifically targeted antiviral therapy for HCV (STAT-C molecules achieve rapid viral suppression and very high rapid virological response rates, and improve sustained virological response rates. The attrition rate of agents within this class has been high due to various toxicities. Regardless, several STAT-C molecules are poised to become the standard of care for HCV treatment in the foreseeable future. Optimism must be tempered with concerns related to the rapid development of drug resistance with resulting HCV rebound. Strategies including induction dosing with interferon and ribavirin, use of combination high-potency STAT-C molecules and an intensive emphasis on adherence to HCV antiviral therapy will be critical to the success of this promising advance in HCV therapy.

  1. Control of target gene specificity during metamorphosis by the steroid response gene E93.

    Science.gov (United States)

    Mou, Xiaochun; Duncan, Dianne M; Baehrecke, Eric H; Duncan, Ian

    2012-02-21

    Hormonal control of sexual maturation is a common feature in animal development. A particularly dramatic example is the metamorphosis of insects, in which pulses of the steroid hormone ecdysone drive the wholesale transformation of the larva into an adult. The mechanisms responsible for this transformation are not well understood. Work in Drosophila indicates that the larval and adult forms are patterned by the same underlying sets of developmental regulators, but it is not understood how the same regulators pattern two distinct forms. Recent studies indicate that this ability is facilitated by a global change in the responsiveness of target genes during metamorphosis. Here we show that this shift is controlled in part by the ecdysone-induced transcription factor E93. Although long considered a dedicated regulator of larval cell death, we find that E93 is expressed widely in adult cells at the pupal stage and is required for many patterning processes at this time. To understand the role of E93 in adult patterning, we focused on a simple E93-dependent process, the induction of the Dll gene within bract cells of the pupal leg by EGF receptor signaling. In this system, we show that E93 functions to cause Dll to become responsive to EGF receptor signaling. We demonstrate that E93 is both necessary and sufficient for directing this switch. E93 likely controls the responsiveness of many other target genes because it is required broadly for patterning during metamorphosis. The wide conservation of E93 orthologs suggests that similar mechanisms control life-cycle transitions in other organisms, including vertebrates.

  2. Control of target gene specificity during metamorphosis by the steroid response gene E93.

    Science.gov (United States)

    Mou, Xiaochun; Duncan, Dianne M; Baehrecke, Eric H; Duncan, Ian

    2012-02-21

    Hormonal control of sexual maturation is a common feature in animal development. A particularly dramatic example is the metamorphosis of insects, in which pulses of the steroid hormone ecdysone drive the wholesale transformation of the larva into an adult. The mechanisms responsible for this transformation are not well understood. Work in Drosophila indicates that the larval and adult forms are patterned by the same underlying sets of developmental regulators, but it is not understood how the same regulators pattern two distinct forms. Recent studies indicate that this ability is facilitated by a global change in the responsiveness of target genes during metamorphosis. Here we show that this shift is controlled in part by the ecdysone-induced transcription factor E93. Although long considered a dedicated regulator of larval cell death, we find that E93 is expressed widely in adult cells at the pupal stage and is required for many patterning processes at this time. To understand the role of E93 in adult patterning, we focused on a simple E93-dependent process, the induction of the Dll gene within bract cells of the pupal leg by EGF receptor signaling. In this system, we show that E93 functions to cause Dll to become responsive to EGF receptor signaling. We demonstrate that E93 is both necessary and sufficient for directing this switch. E93 likely controls the responsiveness of many other target genes because it is required broadly for patterning during metamorphosis. The wide conservation of E93 orthologs suggests that similar mechanisms control life-cycle transitions in other organisms, including vertebrates. PMID:22308414

  3. Selective killing of cancer cells by small molecules targeting heat shock stress response.

    Science.gov (United States)

    Zhang, Daniel; Zhang, Bin

    2016-09-30

    HSF1 heat shock response has emerged as a valuable non-oncogenetic intervention point in targeted cancer therapy. Current reporter based high throughput screening has led to the discovery of several compounds or chemotypes that are effective in the growth inhibition of multiple cancer cell lines and relevant animal tumor models. However, some intrinsic limitations of reporter based assays can potentially lead to biased results. Using a previously validated high content image based assay, we performed a phenotypic screen targeting HSF1 heat shock pathway with a chemically diversified library of over 100,000 compounds. Several novel functional inhibitors of HSF1 pathway were identified with different chemotypes. Western blot analysis confirmed that selective compounds inhibit phosphorylation of HSF1, followed by reduced expression of HSP proteins. Moreover, HeLa cells stably transfected with HSF1 shRNA were more resistant to the compound treatment under lethal temperature than cells containing HSF1, validating HSF1 dependent mechanism of action. These compounds demonstrate nanomolar potency toward multiple cancer cell lines with relatively low cytotoxicity to normal cells. Further SAR and target identification study will pave the way for the potential development of next generation anticancer drugs. PMID:27553278

  4. Recognizing subsurface target responses in ground penetrating radar data using convolutional neural networks

    Science.gov (United States)

    Sakaguchi, Rayn T.; Morton, Kenneth D.; Collins, Leslie M.; Torrione, Peter A.

    2015-05-01

    Improved performance in the discrimination of buried threats using Ground Penetrating Radar (GPR) data has recently been achieved using features developed for applications in computer vision. These features, designed to characterize local shape information in images, have been utilized to recognize patches that contain a target signature in two-dimensional slices of GPR data. While these adapted features perform very well in this GPR application, they were not designed to specifically differentiate between target responses and background GPR data. One option for developing a feature specifically designed for target differentiation is to manually design a feature extractor based on the physics of GPR image formation. However, as seen in the historical progression of computer vision features, this is not a trivial task. Instead, this research evaluates the use of convolutional neural networks (CNNs) applied to two-dimensional GPR data. The benefit of using a CNN is that features extracted from the data are a learned parameter of the system. This has allowed CNN implementations to achieve state of the art performance across a variety of data types, including visual images, without the need for expert designed features. However, the implementation of a CNN must be done carefully for each application as network parameters can cause performance to vary widely. This paper presents results from using CNNs for object detection in GPR data and discusses proper parameter settings and other considerations.

  5. Acidic Tumor pH-Responsive Nanophotomedicine for Targeted Photodynamic Cancer Therapy

    Directory of Open Access Journals (Sweden)

    Wooram Park

    2016-01-01

    Full Text Available An acidic tumor pH-responsive nanophotomedicine (pH-NanoPM for targeted photodynamic therapy (PDT was demonstrated herein. The pH-NanoPM was prepared with a size of ~110 nm by self-assembly of a pH-responsive polymeric photosensitizer (pH-PPS consisting of pH-cleavable methoxypolyethylene glycol (pH-C-mPEG. Because the pH-C-mPEG can be detached from the nanoparticles by hydrolysis of the benzoic-imine group at the pH of an acidic tumor (~6.5, the particle size and surface charge of the pH-NanoPM were changed along with the environmental pH condition. After detachment of the pH-C-mPEG, the pH-NanoPM particles became positively charged (+18.67±1.95 mV due to exposure of primary amine groups and decreased to a size of ~40 nm. From in vitro cellular experiments with HeLa human cervical cancer cells, the pH-NanoPM exhibited enhanced cellular internalization at acidic tumor pH compared to normal pH, which led to a significant cancer cell killing effect. These results suggest that this system has the potential to be used as a new class of nanophotomedicine for targeted photodynamic cancer therapy.

  6. Adjoint affine fusion and tadpoles

    Science.gov (United States)

    Urichuk, Andrew; Walton, Mark A.

    2016-06-01

    We study affine fusion with the adjoint representation. For simple Lie algebras, elementary and universal formulas determine the decomposition of a tensor product of an integrable highest-weight representation with the adjoint representation. Using the (refined) affine depth rule, we prove that equally striking results apply to adjoint affine fusion. For diagonal fusion, a coefficient equals the number of nonzero Dynkin labels of the relevant affine highest weight, minus 1. A nice lattice-polytope interpretation follows and allows the straightforward calculation of the genus-1 1-point adjoint Verlinde dimension, the adjoint affine fusion tadpole. Explicit formulas, (piecewise) polynomial in the level, are written for the adjoint tadpoles of all classical Lie algebras. We show that off-diagonal adjoint affine fusion is obtained from the corresponding tensor product by simply dropping non-dominant representations.

  7. Adjoint affine fusion and tadpoles

    CERN Document Server

    Urichuk, Andrew

    2016-01-01

    We study affine fusion with the adjoint representation. For simple Lie algebras, elementary and universal formulas determine the decomposition of a tensor product of an integrable highest-weight representation with the adjoint representation. Using the (refined) affine depth rule, we prove that equally striking results apply to adjoint affine fusion. For diagonal fusion, a coefficient equals the number of nonzero Dynkin labels of the relevant affine highest weight, minus 1. A nice lattice-polytope interpretation follows, and allows the straightforward calculation of the genus-1 1-point adjoint Verlinde dimension, the adjoint affine fusion tadpole. Explicit formulas, (piecewise) polynomial in the level, are written for the adjoint tadpoles of all classical Lie algebras. We show that off-diagonal adjoint affine fusion is obtained from the corresponding tensor product by simply dropping non-dominant representations.

  8. Consumer affinity for foreign countries: construct development, buying behavior consequences and animosity contrasts

    OpenAIRE

    Nes, Erik B.; Yelkur, Rama; Silkoset, Ragnhild

    2014-01-01

    Purpose: Our purpose is to extend affinity theory in construct domain, scale development, model testing and by discerning affinity and animosity. Design/methodology/approach: We carry out exploratory and empirical research in order to explore the domain and to test the factor structure and the hypotheses through confirmatory analysis. Findings: We find (1) four target country affinity dimensions, (2) consumer affinity impacts micro country image, buying intentions and actual product own...

  9. Correction: Stimuli-responsive magnetic nanoparticles for tumor-targeted bimodal imaging and photodynamic/hyperthermia combination therapy.

    Science.gov (United States)

    Kim, Kyoung Sub; Kim, Jiyoung; Lee, Joo Young; Matsuda, Shofu; Hideshima, Sho; Mori, Yasurou; Osaka, Tetsuya; Na, Kun

    2016-07-01

    Correction for 'Stimuli-responsive magnetic nanoparticles for tumor-targeted bimodal imaging and photodynamic/hyperthermia combination therapy' by Kyoung Sub Kim, et al., Nanoscale, 2016, DOI: 10.1039/c6nr02273a. PMID:27300478

  10. Correction: Stimuli-responsive magnetic nanoparticles for tumor-targeted bimodal imaging and photodynamic/hyperthermia combination therapy

    Science.gov (United States)

    Kim, Kyoung Sub; Kim, Jiyoung; Lee, Joo Young; Matsuda, Shofu; Hideshima, Sho; Mori, Yasurou; Osaka, Tetsuya; Na, Kun

    2016-06-01

    Correction for `Stimuli-responsive magnetic nanoparticles for tumor-targeted bimodal imaging and photodynamic/hyperthermia combination therapy' by Kyoung Sub Kim, et al., Nanoscale, 2016, DOI: 10.1039/c6nr02273a.

  11. Affinity Monolith-Integrated Microchips for Protein Purification and Concentration.

    Science.gov (United States)

    Gao, Changlu; Sun, Xiuhua; Wang, Huaixin; Qiao, Wei; Hu, Bo

    2016-01-01

    Affinity chromatography is a valuable method to purify and concentrate minute amount of proteins. Monoliths with epoxy groups for affinity immobilization were prepared by direct in-situ photopolymerization of glycidyl methacrylate and ethylene glycol dimethacrylate in porogenic solvents consisting of 1-dodecanol and cyclohexanol. By integrating affinity monoliths onto a microfluidic system, targeted biomolecules can be captured and retained on affinity column, while other biomolecules having no specific interactions toward the immobilized ligands flow through the microchannel. Therefore, proteins which remain on the affinity column are purified and concentrated, and then eluted by appropriate solutions and finally, separated by microchip capillary electrophoresis. This integrated microfluidic device has been applied to the purification and separation of specific proteins (FITC-labeled human serum albumin and IgG) in a mixture.

  12. Detection of protein-protein interactions using tandem affinity purification.

    Science.gov (United States)

    Goodfellow, Ian; Bailey, Dalan

    2014-01-01

    Tandem affinity purification (TAP) is an invaluable technique for identifying interaction partners for an affinity tagged bait protein. The approach relies on the fusion of dual tags to the bait before separate rounds of affinity purification and precipitation. Frequently two specific elution steps are also performed to increase the specificity of the overall technique. In the method detailed here, the two tags used are protein G and a short streptavidin binding peptide; however, many variations can be employed. In our example the tags are separated by a cleavable tobacco etch virus protease target sequence, allowing for specific elution after the first round of affinity purification. Proteins isolated after the final elution step in this process are concentrated before being identified by mass spectrometry. The use of dual affinity tags and specific elution in this technique dramatically increases both the specificity and stringency of the pull-downs, ensuring a low level of background nonspecific interactions.

  13. PAS-1, a protein affinity purified from Ascaris suum worms, maintains the ability to modulate the immune response to a bystander antigen.

    Science.gov (United States)

    Oshiro, Telma M; Enobe, Cristina S; Araújo, Cláudia A; Macedo, Mahasti S; Macedo-Soares, Maria Fernanda

    2006-04-01

    Helminth infections and parasite components have potent immunomodulatory effects on a host's immune system. In the present study, we investigated the effect of PAS-1, a protein component of Ascaris suum adult worms recognized by a monoclonal antibody (MAIP-1), on humoral and cell-mediated responses to a bystander antigen (ovalbumin [OVA]). MAIP-1 recognized only one of the three polypeptide chains of PAS-1, but neutralized the suppressive effect of the whole worm extract on OVA-specific antibody production. PAS-1 inhibited antibody production against a T-cell-dependent, but not a T-cell-independent, antigen in a dose-dependent way. IgM, IgG1, IgG2b, and also IgE and anaphylactic IgG1 levels were downregulated. In addition, PAS-1 inhibited OVA-specific delayed type hypersensitivity reactions in the footpad of mice, showing a potent immunosuppressive activity on both Th1 and Th2 responses that seems to be mediated by the induction of large amounts of IL-10 and IL-4. Indeed, PAS-1-specific spleen cells secreted sevenfold more IL-10 and threefold more IL-4 than OVA-specific cells in response to in vitro restimulation with the respective antigens. In conclusion, we showed that PAS-1, a single protein component from A. suum, maintains all its immunosuppressive properties. PMID:16519731

  14. High- and low-affinity binding of S-citalopram to the human serotonin transporter mutated at 20 putatively important amino acid positions

    DEFF Research Database (Denmark)

    Plenge, Per; Wiborg, Ove

    2005-01-01

    The serotonin transporter (SERT) is responsible for terminating or modulating the action of serotonin released from the presynaptic neuron and is the major target for most antidepressants including the tricyclic antidepressants and the selective serotonin uptake inhibitors. Two binding sites...... for uptake inhibitors and serotonin (5-HT) have been found on SERT. At one site, uptake inhibitors bind with high-affinity to SERT, thereby blocking the uptake of 5-HT. The other site is a low-affinity allosteric site, which influences the dissociation of uptake inhibitors, such as imipramine, paroxetine......, and citalopram from the first site, when occupied by 5-HT and a few uptake inhibitors like paroxetine and citalopram. In this study, the connection between the high-affinity binding site and the allosteric affinity-modulating site was investigated by introducing 20 single amino acid substitutions into positions...

  15. The Leishmania infantum PUF proteins are targets of the humoral response during visceral leishmaniasis

    Directory of Open Access Journals (Sweden)

    Requena Jose M

    2010-01-01

    Full Text Available Abstract Background RNA-binding proteins of the PUF family share a conserved domain consisting of tandemly repeated 36-40 amino acid motifs (typically eight known as Puf repeats. Proteins containing tandem repeats are often dominant targets of humoral responses during infectious diseases. Thus, we considered of interest to analyze whether Leishmania PUF proteins result antigenic during visceral leishmaniasis (VL. Findings Here, employing whole-genome databases, we report the composition, and structural features, of the PUF family in Leishmania infantum. Additionally, the 10 genes of the L. infantum PUF family were cloned and used to express the Leishmania PUFs in bacteria as recombinant proteins. Finally, the antigenicity of these PUF proteins was evaluated by determining levels of specific antibodies in sera from experimentally infected hamsters. The Leishmania PUFs were all recognized by the sera, even though with different degree of reactivity and/or frequency of recognition. The reactivity of hamster sera against recombinant LiPUF1 and LiPUF2 was particularly prominent, and these proteins were subsequently assayed against sera from human patients. High antibody responses against rLiPUF1 and rLiPUF2 were found in sera from VL patients, but these proteins resulted also recognized by sera from Chagas' disease patients. Conclusion Our results suggest that Leishmania PUFs are targets of the humoral response during L. infantum infection and may represent candidates for serodiagnosis and/or vaccine reagents; however, it should be kept in mind the cross-reactivity of LiPUFs with antibodies induced against other trypanosomatids such as Trypanosoma cruzi.

  16. Regulation of microRNA-155 in atherosclerotic inflammatory responses by targeting MAP3K10.

    Directory of Open Access Journals (Sweden)

    Jianhua Zhu

    Full Text Available AIMS: Accumulating evidence suggest that numerous microRNAs (miRNAs play important roles in cell proliferation, apoptosis, and differentiation, as well as various diseases that accompany inflammatory responses. Inflammation is known to be a major contributor to atherogenesis. Previous studies provide promising evidence in support of the role of miRNAs in cardiovascular disease. However, mechanistic data on these small molecules in atherosclerosis (AS are still missing. The present study aims to investigate the potential role of miRNAs in AS. METHODS AND RESULTS: The miRNA transcriptase was verified by TaqMan real-time polymerase chain reaction assay. Thoracic aorta samples were obtained from Apolipoprotein E knockout mice, and plasma samples were from coronary artery disease (CAD patients. The results showed that the miR-155 level was the most significantly elevated both in AS mice and CAD patients relative to the normal control. The functional role of miR-155 in the atherosclerotic path physiological process was also observed in vivo and in vitro. The observations suggested that miR-155 is a part of a negative feedback loop, which down-modulates inflammatory cytokine production and decreases AS progression. miR-155 was also found to mediate the inflammatory response and mitogen-activated protein kinase (MAPK pathway by targeting mitogen-activated protein kinase kinase kinase 10. CONCLUSIONS: miR-155 contributes to the prevention of AS development and progression. It may also be involved in the posttranscriptional regulation of the inflammatory response and MAPK pathway by targeting mitogen-activated protein kinase kinase kinase 10.

  17. Targeting the cell stress response of Plasmodium falciparum to overcome artemisinin resistance.

    Science.gov (United States)

    Dogovski, Con; Xie, Stanley C; Burgio, Gaetan; Bridgford, Jess; Mok, Sachel; McCaw, James M; Chotivanich, Kesinee; Kenny, Shannon; Gnädig, Nina; Straimer, Judith; Bozdech, Zbynek; Fidock, David A; Simpson, Julie A; Dondorp, Arjen M; Foote, Simon; Klonis, Nectarios; Tilley, Leann

    2015-04-01

    Successful control of falciparum malaria depends greatly on treatment with artemisinin combination therapies. Thus, reports that resistance to artemisinins (ARTs) has emerged, and that the prevalence of this resistance is increasing, are alarming. ART resistance has recently been linked to mutations in the K13 propeller protein. We undertook a detailed kinetic analysis of the drug responses of K13 wild-type and mutant isolates of Plasmodium falciparum sourced from a region in Cambodia (Pailin). We demonstrate that ART treatment induces growth retardation and an accumulation of ubiquitinated proteins, indicative of a cellular stress response that engages the ubiquitin/proteasome system. We show that resistant parasites exhibit lower levels of ubiquitinated proteins and delayed onset of cell death, indicating an enhanced cell stress response. We found that the stress response can be targeted by inhibiting the proteasome. Accordingly, clinically used proteasome inhibitors strongly synergize ART activity against both sensitive and resistant parasites, including isogenic lines expressing mutant or wild-type K13. Synergy is also observed against Plasmodium berghei in vivo. We developed a detailed model of parasite responses that enables us to infer, for the first time, in vivo parasite clearance profiles from in vitro assessments of ART sensitivity. We provide evidence that the clinical marker of resistance (delayed parasite clearance) is an indirect measure of drug efficacy because of the persistence of unviable parasites with unchanged morphology in the circulation, and we suggest alternative approaches for the direct measurement of viability. Our model predicts that extending current three-day ART treatment courses to four days, or splitting the doses, will efficiently clear resistant parasite infections. This work provides a rationale for improving the detection of ART resistance in the field and for treatment strategies that can be employed in areas with ART

  18. Magnetically-responsive silica-gold nanobowls for targeted delivery and SERS-based sensing

    Science.gov (United States)

    Mo, Alexander H.; Landon, Preston B.; Gomez, Karla Santacruz; Kang, Heemin; Lee, Joon; Zhang, Chen; Janetanakit, Woraphong; Sant, Vrinda; Lu, Tianyu; Colburn, David A.; Akkiraju, Siddhartha; Dossou, Samuel; Cao, Yue; Lee, Kuo-Fen; Varghese, Shyni; Glinsky, Gennadi; Lal, Ratnesh

    2016-06-01

    Composite colloidal structures with multi-functional properties have wide applications in targeted delivery of therapeutics and imaging contrast molecules and high-throughput molecular bio-sensing. We have constructed a multifunctional composite magnetic nanobowl using the bottom-up approach on an asymmetric silica/polystyrene Janus template consisting of a silica shell around a partially exposed polystyrene core. The nanobowl consists of a silica bowl and a gold exterior shell with iron oxide magnetic nanoparticles sandwiched between the silica and gold shells. The nanobowls were characterized by electron microscopy, atomic force microscopy, magnetometry, vis-NIR and FTIR spectroscopy. Magnetically vectored transport of these nanobowls was ascertained by time-lapsed imaging of their flow in fluid through a porous hydrogel under a defined magnetic field. These magnetically-responsive nanobowls show distinct surface enhanced Raman spectroscopy (SERS) imaging capability. The PEGylated magnetically-responsive nanobowls show size-dependent cellular uptake in vitro.Composite colloidal structures with multi-functional properties have wide applications in targeted delivery of therapeutics and imaging contrast molecules and high-throughput molecular bio-sensing. We have constructed a multifunctional composite magnetic nanobowl using the bottom-up approach on an asymmetric silica/polystyrene Janus template consisting of a silica shell around a partially exposed polystyrene core. The nanobowl consists of a silica bowl and a gold exterior shell with iron oxide magnetic nanoparticles sandwiched between the silica and gold shells. The nanobowls were characterized by electron microscopy, atomic force microscopy, magnetometry, vis-NIR and FTIR spectroscopy. Magnetically vectored transport of these nanobowls was ascertained by time-lapsed imaging of their flow in fluid through a porous hydrogel under a defined magnetic field. These magnetically-responsive nanobowls show distinct

  19. Immune response to functionalized mesoporous silica nanoparticles for targeted drug delivery.

    Science.gov (United States)

    Heidegger, Simon; Gössl, Dorothée; Schmidt, Alexandra; Niedermayer, Stefan; Argyo, Christian; Endres, Stefan; Bein, Thomas; Bourquin, Carole

    2016-01-14

    Multifunctional mesoporous silica nanoparticles (MSN) have attracted substantial attention with regard to their high potential for targeted drug delivery. For future clinical applications it is crucial to address safety concerns and understand the potential immunotoxicity of these nanoparticles. In this study, we assess the biocompatibility and functionality of multifunctional MSN in freshly isolated, primary murine immune cells. We show that the functionalized silica nanoparticles are rapidly and efficiently taken up into the endosomal compartment by specialized antigen-presenting cells such as dendritic cells. The silica nanoparticles showed a favorable toxicity profile and did not affect the viability of primary immune cells from the spleen in relevant concentrations. Cargo-free MSN induced only very low immune responses in primary cells as determined by surface expression of activation markers and release of pro-inflammatory cytokines such as Interleukin-6, -12 and -1β. In contrast, when surface-functionalized MSN with a pH-responsive polymer capping were loaded with an immune-activating drug, the synthetic Toll-like receptor 7 agonist R848, a strong immune response was provoked. We thus demonstrate that MSN represent an efficient drug delivery vehicle to primary immune cells that is both non-toxic and non-inflammagenic, which is a prerequisite for the use of these particles in biomedical applications. PMID:26659601

  20. A Data Mining-Based Response Model for Target Selection in Direct Marketing

    Directory of Open Access Journals (Sweden)

    Eniafe Festus Ayetiran

    2012-02-01

    Full Text Available Identifying customers who are more likely to respond to new product offers is an important issue in direct marketing. In direct marketing, data mining has been used extensively to identify potential customers for a new product (target selection. Using historical purchase data, a predictive response model with data mining techniques was developed to predict a probability that a customer in Ebedi Microfinance bank will respond to a promotion or an offer. To achieve this purpose, a predictive response model using customers’ historical purchase data was built with data mining techniques. The data were stored in a data warehouse to serve as management decision support system. The response model was built from customers’ historic purchases and demographic dataset.Bayesian algorithm precisely Naïve Bayes algorithm was employed in constructing the classifier system. Both filter and wrapper feature selection techniques were employed in determining inputs to the model.The results obtained shows that Ebedi Microfinance bank can plan effective marketing of their products and services by obtaining a guiding report on the status of their customers which will go a long way in assisting management in saving significant amount of money that could have been spent on wasteful promotional campaigns.

  1. Target-responsive aptamer release from manganese dioxide nanosheets for electrochemical sensing of cocaine with target recycling amplification.

    Science.gov (United States)

    Chen, Zongbao; Lu, Minghua

    2016-11-01

    A novel electrochemical sensing platform based on manganese dioxide (MnO2) nanosheets was developed for sensitive screening of target cocaine with the signal amplification. Ferrocene-labeled cocaine aptamers were initially immobilized onto MnO2 nanosheets-modified screen-printed carbon electrode because of π-stacking interaction between nucleobases and nanosheets. The immobilized ferrocene-aptamer activated the electrical contact with the electrode, thereby resulting in the sensor circuit to switch on. Upon target cocaine introduction, the analyte reacted with the aptamer and caused the dissociation of ferrocene-aptamer from the electrode, thus giving rise to the detection circuit to switch off. The released aptamer was cleaved by DNase I with target recycling. Under optimal conditions, the decreasing percentage of the electronic signal relative to background current increased with the increasing cocaine concentration in the dynamic range of 0.1-20nM, and the detection limit was 32pM. The reproducibility, selectivity and method accuracy were acceptable. Importantly, this concept offers promise for rapid, simple, and cost-effective analysis of cocaine biological samples without the needs of sample separation and multiple washing steps. PMID:27591636

  2. Enhanced radiation response in radioresistant MCF-7 cells by targeting peroxiredoxin II

    Directory of Open Access Journals (Sweden)

    Diaz AJG

    2013-10-01

    Full Text Available Anthony Joseph Gomez Diaz,1 Daniel Tamae,2 Yun Yen,3 JianJian Li,4 Tieli Wang1 1Department of Chemistry and Biochemistry, California State University at Dominguez Hills, Carson, CA, 2Center of Excellence in Environmental Toxicology, Department of Pharmacology, University of Pennsylvania, Philadelphia, PA, 3Department of Clinical and Molecular Pharmacology, Beckman Research Institute of City of Hope National Medical Center, Duarte, CA, 4Department of Radiation Oncology, University of California Davis, Sacramento, CA, USA Abstract: In our previous study, we identified that a protein target, peroxiredoxin II (PrxII, is overexpressed in radioresistant MCF+FIR3 breast-cancer cells and found that its expression and function is associated with breast-cancer radiation sensitivity or resistance. Small interference RNA (siRNA targeting PrxII gene expression was able to sensitize MCF+FIR3 radioresistant breast-cancer cells to ionizing radiation. The major focus of this work was to investigate how the radiation response of MCF+FIR3 radioresistant cells was affected by the siRNA that inhibits PrxII gene expression. Our results, presented here, show that silencing PrxII gene expression increased cellular toxicity by altering cellular thiol status, inhibiting Ca2+ efflux from the cells, and perturbing the intracellular Ca2+ homeostasis. By combining radiotherapy and siRNA technology, we hope to develop new therapeutic strategies that may have potential to enhance the efficacy of chemotherapeutic agents due to this technology's property of targeting to specific cancer-related genes. Keywords: siRNA, PrxII, radiation resistance, Ca2+, MCF+FIR3

  3. Immune response to functionalized mesoporous silica nanoparticles for targeted drug delivery

    CERN Document Server

    Heidegger, S; Schmidt, A; Gößl, D; Argyo, C; Endres, S; Bein, T; Bourquin, C

    2015-01-01

    Multifunctional mesoporous silica nanoparticles (MSN) have attracted substantial attention with regard to their high potential for targeted drug delivery. For future clinical applications it is crucial to address safety concerns and understand the potential immunotoxicity of these nanoparticles. In this study, we assess the biocompatibility and functionality of multifunctional MSN in freshly isolated, primary murine immune cells. We show that the functionalized silica nanoparticles are rapidly and efficiently taken up into the endosomal compartment by specialized antigen-presenting cells such as dendritic cells. The silica nanoparticles showed a favorable toxicity profile and did not affect the viability of primary immune cells from the spleen in relevant concentrations. Cargo-free MSN induced only very low immune responses in primary cells as determined by surface expression of activation markers and release of pro-inflammatory cytokines such as Interleukin-6, -12 and -1\\beta. In contrast, when surface-funct...

  4. Targeting the microenvironment of pancreatic cancer: overcoming treatment barriers and improving local immune responses.

    Science.gov (United States)

    Strauss, J; Alewine, C; Figg, W D; Duffy, A

    2016-07-01

    Historically, patients diagnosed with metastatic pancreatic cancer have faced a grim prognosis. The survival benefit seen with systemic chemotherapies and even combinations thereof have been disappointing. However, growing data suggest that the microenvironment of pancreatic cancer may be contributing to this poor prognosis. This microenvironment has a dense fibrotic stroma, and is hypoxic and highly immunosuppressive, all of which pose barriers to treatment. Newer strategies looking to disrupt the fibrotic stroma, target hypoxic areas, and improve local immune responses in the tumor microenvironment are currently undergoing clinical evaluation and seem to offer great promise. In addition to these therapies, preclinical work evaluating novel cytotoxic agents including nanoparticles has also been encouraging. While much research still needs to be done, these strategies offer new hope for patients with pancreatic cancer. PMID:26661112

  5. Response Simulation of a Micro Reinforced Concrete Target Under Ballistic Impact

    Science.gov (United States)

    Mohan, V.; Rajasankar, J.; Iyer, N. R.

    2014-05-01

    The response of concrete structures subjected to impact loading has received extensive attention in both civil and military applications. Research on improving the shock resistance of concrete has led to the development of cementitious composites. Micro Reinforced Concrete (MRC), a type of cementitious composite, is a concrete matrix embedded with multilayered steel wire meshes. This paper presents 3D hydrocode simulations of MRC panels subjected to impact under a ballistic range. A finite element model based on Lagrange formulation is used to represent both a 300 mm × 300 mm × 100 mm target with 30 layers of wire mesh and a 5.56 × 45 mm projectile in simulations. Penetration depth and damage patterns of the MRC mesh cement composite panel are numerically compared with those of the field experiment. The results show a relatively good agreement.

  6. Reactive oxygen species-responsive protein modification and its intracellular delivery for targeted cancer therapy.

    Science.gov (United States)

    Wang, Ming; Sun, Shuo; Neufeld, Caleb I; Perez-Ramirez, Bernardo; Xu, Qiaobing

    2014-12-01

    Herein we report a convenient chemical approach to reversibly modulate protein (RNase A) function and develop a protein that is responsive to reactive oxygen species (ROS) for targeted cancer therapy. The conjugation of RNase A with 4-nitrophenyl 4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl) benzyl carbonate (NBC) blocks protein lysine and temporarily deactivates the protein. However, the treatment of RNase A-NBC with hydrogen peroxide (one major intracellular ROS) efficiently cleaves the NBC conjugation and restores the RNase A activity. Thus, RNase A-NBC can be reactivated inside tumor cells by high levels of intracellular ROS, thereby restoring the cytotoxicity of RNase A for cancer therapy. Due to higher ROS levels inside tumor cells compared to healthy cells, and the resulting different levels of RNase A-NBC reactivation, RNase A-NBC shows a significant specific cytotoxicity against tumor cells.

  7. Lineup identification accuracy: The effects of alcohol, target presence, confidence ratings, and response time

    Directory of Open Access Journals (Sweden)

    Wendy Kneller

    2016-01-01

    Full Text Available Despite the intoxication of many eyewitnesses at crime scenes, only four published studies to date have investigated the effects of alcohol intoxication on eyewitness identification performance. While one found intoxication significantly increased false identification rates from target absent showups, three found no such effect using the more traditional lineup procedure. The present study sought to further explore the effects of alcohol intoxication on identification performance and examine whether accurate decisions from intoxicated witnesses could be postdicted by confidence and response times. One hundred and twenty participants engaged in a study examining the effects of intoxication (control, placebo, and mild intoxication and target presence on identification performance. Participants viewed a simultaneous lineup one week after watching a mock crime video of a man attempting to steal cars. Ethanol intoxication (0.6 ml/kg was found to make no significant difference to identification accuracy and such identifications from intoxicated individuals were made no less confidently or slowly than those from sober witnesses. These results are discussed with respect to the previous research examining intoxicated witness identification accuracy and the misconceptions the criminal justice system holds about the accuracy of such witnesses.

  8. Sickle erythrocytes target cytotoxics to hypoxic tumor microvessels and potentiate a tumoricidal response.

    Directory of Open Access Journals (Sweden)

    David S Terman

    Full Text Available Resistance of hypoxic solid tumor niches to chemotherapy and radiotherapy remains a major scientific challenge that calls for conceptually new approaches. Here we exploit a hitherto unrecognized ability of sickled erythrocytes (SSRBCs but not normal RBCs (NLRBCs to selectively target hypoxic tumor vascular microenviroment and induce diffuse vaso-occlusion. Within minutes after injection SSRBCs, but not NLRBCs, home and adhere to hypoxic 4T1 tumor vasculature with hemoglobin saturation levels at or below 10% that are distributed over 70% of the tumor space. The bound SSRBCs thereupon form microaggregates that obstruct/occlude up to 88% of tumor microvessels. Importantly, SSRBCs, but not normal RBCs, combined with exogenous prooxidant zinc protoporphyrin (ZnPP induce a potent tumoricidal response via a mutual potentiating mechanism. In a clonogenic tumor cell survival assay, SSRBC surrogate hemin, along with H(2O(2 and ZnPP demonstrate a similar mutual potentiation and tumoricidal effect. In contrast to existing treatments directed only to the hypoxic tumor cell, the present approach targets the hypoxic tumor vascular environment and induces injury to both tumor microvessels and tumor cells using intrinsic SSRBC-derived oxidants and locally generated ROS. Thus, the SSRBC appears to be a potent new tool for treatment of hypoxic solid tumors, which are notable for their resistance to existing cancer treatments.

  9. Tumor responsive targeted multifunctional nanosystems for cancer imaging, chemo- and siRNA therapy

    Science.gov (United States)

    Savla, Ronak

    Cancer is one of the most insidious diseases. Compromising of over 100 different types and sharing the unifying factors of uncontrolled growth and metastasis, unmet clinical needs in terms of cancer diagnosis and treatment continue to exist. It is widely accepted that most forms of cancer are treatable or even curable if detected before widespread metastasis occurs. Nearly a quarter of deaths in the United States is the result of cancer and it only trails heart disease in terms of annual mortality. Surgery, chemotherapy, and radiation therapy are the primary treatment modalities for cancer. Research in these procedures has resulted in substantial benefits for cancer patients, but there is still room for an improvement. However, a time has been reached at which it appears that the benefits from these modalities have been reached the maximum. Therefore, it is vital to develop new strategies for the diagnosis and treatment of cancer. The field of nanotechnology is concerned with structures in the nanometer size range and holds the potential to drastically impact and improve the lives of patients suffering from cancer. Not only can nanotechnology improve current methods of diagnosis and treatment, it has a possibility of introducing newer and better modalities. The overall purpose of this work is to develop novel nanotechnology-based methodologies for the diagnosis and treatment of various forms of cancers. The first aim of the project is the development of a multifunctional targeted nanosystem for the delivery of siRNA to overcome drug resistance. The second aspect is the synthesis of a quantum dot-based delivery system that releases drug in response to pH changes. The third aim is the development of a targeted, tumor environment responsive magnetic resonance nanoparticle contrast agent coupled with a nanoparticle-based treatment.

  10. miR-204 targets Bcl-2 expression and enhances responsiveness of gastric cancer

    Science.gov (United States)

    Sacconi, A; Biagioni, F; Canu, V; Mori, F; Di Benedetto, A; Lorenzon, L; Ercolani, C; Di Agostino, S; Cambria, A M; Germoni, S; Grasso, G; Blandino, R; Panebianco, V; Ziparo, V; Federici, O; Muti, P; Strano, S; Carboni, F; Mottolese, M; Diodoro, M; Pescarmona, E; Garofalo, A; Blandino, G

    2012-01-01

    Micro RNAs (miRs) are small non-coding RNAs aberrantly expressed in human tumors. Here, we aim to identify miRs whose deregulated expression leads to the activation of oncogenic pathways in human gastric cancers (GCs). Thirty nine out of 123 tumoral and matched uninvolved peritumoral gastric specimens from three independent European subsets of patients were analyzed for the expression of 851 human miRs using Agilent Platform. The remaining 84 samples were used to validate miRs differentially expressed between tumoral and matched peritumoral specimens by qPCR. miR-204 falls into a group of eight miRs differentially expressed between tumoral and peritumoral samples. Downregulation of miR-204 has prognostic value and correlates with increased staining of Bcl-2 protein in tumoral specimens. Ectopic expression of miR-204 inhibited colony forming ability, migration and tumor engraftment of GC cells. miR-204 targeted Bcl-2 messenger RNA and increased responsiveness of GC cells to 5-fluorouracil and oxaliplatin treatment. Ectopic expression of Bcl-2 protein counteracted miR-204 pro-apoptotic activity in response to 5-fluorouracil. Altogether, these findings suggest that modulation of aberrant expression of miR-204, which in turn releases oncogenic Bcl-2 protein activity might hold promise for preventive and therapeutic strategies of GC. PMID:23152059

  11. pH-Sensitive stimulus-responsive nanocarriers for targeted delivery of therapeutic agents.

    Science.gov (United States)

    Karimi, Mahdi; Eslami, Masoud; Sahandi-Zangabad, Parham; Mirab, Fereshteh; Farajisafiloo, Negar; Shafaei, Zahra; Ghosh, Deepanjan; Bozorgomid, Mahnaz; Dashkhaneh, Fariba; Hamblin, Michael R

    2016-09-01

    In recent years miscellaneous smart micro/nanosystems that respond to various exogenous/endogenous stimuli including temperature, magnetic/electric field, mechanical force, ultrasound/light irradiation, redox potentials, and biomolecule concentration have been developed for targeted delivery and release of encapsulated therapeutic agents such as drugs, genes, proteins, and metal ions specifically at their required site of action. Owing to physiological differences between malignant and normal cells, or between tumors and normal tissues, pH-sensitive nanosystems represent promising smart delivery vehicles for transport and delivery of anticancer agents. Furthermore, pH-sensitive systems possess applications in delivery of metal ions and biomolecules such as proteins, insulin, etc., as well as co-delivery of cargos, dual pH-sensitive nanocarriers, dual/multi stimuli-responsive nanosystems, and even in the search for new solutions for therapy of diseases such as Alzheimer's. In order to design an optimized system, it is necessary to understand the various pH-responsive micro/nanoparticles and the different mechanisms of pH-sensitive drug release. This should be accompanied by an assessment of the theoretical and practical challenges in the design and use of these carriers. WIREs Nanomed Nanobiotechnol 2016, 8:696-716. doi: 10.1002/wnan.1389 For further resources related to this article, please visit the WIREs website. PMID:26762467

  12. Apolipoprotein E-specific innate immune response in astrocytes from targeted replacement mice

    Directory of Open Access Journals (Sweden)

    Montine Thomas J

    2006-04-01

    Full Text Available Abstract Background Inheritance of the three different alleles of the human apolipoprotein (apo E gene (APOE are associated with varying risk or clinical outcome from a variety of neurologic diseases. ApoE isoform-specific modulation of several pathogenic processes, in addition to amyloid β metabolism in Alzheimer's disease, have been proposed: one of these is innate immune response by glia. Previously we have shown that primary microglia cultures from targeted replacement (TR APOE mice have apoE isoform-dependent innate immune activation and paracrine damage to neurons that is greatest with TR by the ε4 allele (TR APOE4 and that derives from p38 mitogen-activated protein kinase (p38MAPK activity. Methods Primary cultures of TR APOE2, TR APOE3 and TR APOE4 astrocytes were stimulated with lipopolysaccharide (LPS. ApoE secretion, cytokine production, and nuclear factor-kappa B (NF-κB subunit activity were measured and compared. Results Here we showed that activation of primary astrocytes from TR APOE mice with LPS led to TR APOE-dependent differences in cytokine secretion that were greatest in TR APOE2 and that were associated with differences in NF-κB subunit activity. Conclusion Our results suggest that LPS activation of innate immune response in TR APOE glia results in opposing outcomes from microglia and astrocytes as a result of TR APOE-dependent activation of p38MAPK or NF-κB signaling in these two cell types.

  13. Mechanisms of DNA damage response to targeted irradiation in organotypic 3D skin cultures.

    Directory of Open Access Journals (Sweden)

    Anna Acheva

    Full Text Available DNA damage (caused by direct cellular exposure and bystander signaling and the complex pathways involved in its repair are critical events underpinning cellular and tissue response following radiation exposures. There are limited data addressing the dynamics of DNA damage induction and repair in the skin particularly in areas not directly exposed. Here we investigate the mechanisms regulating DNA damage, repair, intracellular signalling and their impact on premature differentiation and development of inflammatory-like response in the irradiated and surrounding areas of a 3D organotypic skin model. Following localized low-LET irradiation (225 kVp X-rays, low levels of 53BP1 foci were observed in the 3D model (3.8±0.28 foci/Gy/cell with foci persisting and increasing in size up to 48 h post irradiation. In contrast, in cell monolayers 14.2±0.6 foci/Gy/cell and biphasic repair kinetics with repair completed before 24 h was observed. These differences are linked to differences in cellular status with variable level of p21 driving apoptotic signalling in 2D and accelerated differentiation in both the directly irradiated and bystander areas of the 3D model. The signalling pathways utilized by irradiated keratinocytes to induce DNA damage in non-exposed areas of the skin involved the NF-κB transcription factor and its downstream target COX-2.

  14. Intelligently targeted drug delivery and enhanced antitumor effect by gelatinase-responsive nanoparticles.

    Directory of Open Access Journals (Sweden)

    Rutian Li

    Full Text Available AIMS: The matrix metalloproteinase (MMP 2/9, also known as collagenases IV and gelatinases A/B, play a key role in cancer invasion and metastasis. However, the clinical trials of the MMP inhibitors (MMPIs ended up with disappointing results. In this paper, we synthesized a gelatinase-responsive copolymer (mPEG-PCL by inserting a gelatinase cleavable peptide (PVGLIG between mPEG and PCL blocks of mPEG-PCL for anticancer drug delivery to make use of MMP2/9 as an intelligent target for drug delivery. MATERIALS AND METHODS: mPEG-pep-PCL copolymer was synthesized via ring-opening copolymerization and double-amidation. To evaluate whether Nanoparticles (NPs prepared from this copolymer are superior to NPs prepared from mPEG-PCL, NPs prepared from mPEG-PCL copolymer were used as positive control. Docetaxel-loading NPs using mPEG-pep-PCL and mPEG-PCL were prepared by nano-precipitation method, mentioned as Gel-NPs and Con-NPs, respectively. The morphologic changes of the NPs after treatment with gelatinases were observed macroscopically by spectrophotometer and microscopically by transmission electron microscopy (TEM and atomic force microscopy (AFM. The cellular uptake amount and cytotoxicity of Gel-NPs and Con-NPs, respectively, in cell lines with different levels of gelatinase expression were studied. Moreover, the cytotoxicity study on the primary cancer cells isolated from pericardial fluids from a patient with late-stage lung cancer was conducted. RESULTS: The Gel-NPs aggregated in response to gelatinases, which was confirmed macroscopically and microscopically. The cellular uptake amount of Gel-NPs was correlated with the level of gelatinases. The in vitro antitumor effect of Gel-NPs was also correlated with the level of gelatinases and was superior to Taxotere (commercially available docetaxel as well as the Con-NPs. The cytotoxicity study on the primary lung cancer cells also confirmed the effectiveness of Gel-NPs. CONCLUSION: The results in

  15. Immune response to functionalized mesoporous silica nanoparticles for targeted drug delivery

    Science.gov (United States)

    Heidegger, Simon; Gößl, Dorothée; Schmidt, Alexandra; Niedermayer, Stefan; Argyo, Christian; Endres, Stefan; Bein, Thomas; Bourquin, Carole

    2015-12-01

    Multifunctional mesoporous silica nanoparticles (MSN) have attracted substantial attention with regard to their high potential for targeted drug delivery. For future clinical applications it is crucial to address safety concerns and understand the potential immunotoxicity of these nanoparticles. In this study, we assess the biocompatibility and functionality of multifunctional MSN in freshly isolated, primary murine immune cells. We show that the functionalized silica nanoparticles are rapidly and efficiently taken up into the endosomal compartment by specialized antigen-presenting cells such as dendritic cells. The silica nanoparticles showed a favorable toxicity profile and did not affect the viability of primary immune cells from the spleen in relevant concentrations. Cargo-free MSN induced only very low immune responses in primary cells as determined by surface expression of activation markers and release of pro-inflammatory cytokines such as Interleukin-6, -12 and -1β. In contrast, when surface-functionalized MSN with a pH-responsive polymer capping were loaded with an immune-activating drug, the synthetic Toll-like receptor 7 agonist R848, a strong immune response was provoked. We thus demonstrate that MSN represent an efficient drug delivery vehicle to primary immune cells that is both non-toxic and non-inflammagenic, which is a prerequisite for the use of these particles in biomedical applications.Multifunctional mesoporous silica nanoparticles (MSN) have attracted substantial attention with regard to their high potential for targeted drug delivery. For future clinical applications it is crucial to address safety concerns and understand the potential immunotoxicity of these nanoparticles. In this study, we assess the biocompatibility and functionality of multifunctional MSN in freshly isolated, primary murine immune cells. We show that the functionalized silica nanoparticles are rapidly and efficiently taken up into the endosomal compartment by specialized

  16. The application of magnetic force differentiation for the measurement of the affinity of peptide libraries

    International Nuclear Information System (INIS)

    A new method has been developed for measuring the binding affinity of phage displayed peptides and a target protein using magnetic particles. The specific interaction between the phage displayed peptides and the target protein was subject to a force generated by the magnetic particle. The binding affinity was obtained by analyzing the force-bond lifetime

  17. Food and Natural Materials Target Mechanisms to Effectively Regulate Allergic Responses.

    Science.gov (United States)

    Shin, Hee Soon; Shon, Dong-Hwa

    2015-01-01

    An immune hypersensitivity disorder called allergy is caused by diverse allergens entering the body via skin contact, injection, ingestion, and/or inhalation. These allergic responses may develop into allergic disorders, including inflammations such as atopic dermatitis, asthma, anaphylaxis, food allergies, and allergic rhinitis. Several drugs have been developed to treat these allergic disorders; however, long-term intake of these drugs could have adverse effects. As an alternative to these medicines, food and natural materials that ameliorate allergic disorder symptoms without producing any side effects can be consumed. Food and natural materials can effectively regulate successive allergic responses in an allergic chain-reaction mechanism in the following ways: [1] Inhibition of allergen permeation via paracellular diffusion into epithelial cells, [2] suppression of type 2 T-helper (Th) cell-related cytokine production by regulating Th1/Th2 balance, [3] inhibition of pathogenic effector CD4(+) T cell differentiation by inducing regulatory T cells (Treg), and [4] inhibition of degranulation in mast cells. The immunomodulatory effects of food and natural materials on each target mechanism were scientifically verified and shown to alleviate allergic disorder symptoms. Furthermore, consumption of certain food and natural materials such as fenugreek, skullcap, chitin/chitosan, and cheonggukjang as anti-allergics have merits such as safety (no adverse side effects), multiple suppressive effects (as a mixture would contain various components that are active against allergic responses), and ease of consumption when required. These merits and anti-allergic properties of food and natural materials help control various allergic disorders. PMID:26598817

  18. Prenatal nicotine changes the response to postnatal chlorpyrifos: Interactions targeting serotonergic synaptic function and cognition.

    Science.gov (United States)

    Slotkin, Theodore A; Skavicus, Samantha; Levin, Edward D; Seidler, Frederic J

    2015-02-01

    Nicotine and chlorpyrifos are developmental neurotoxicants that target serotonin systems. We examined whether prenatal nicotine exposure alters the subsequent response to chlorpyrifos given postnatally. Pregnant rats received nicotine throughout gestation at 3mg/kg/day, a regimen designed to achieve plasma levels seen in smokers; chlorpyrifos was given to pups on postnatal days (PN) 1-4 at 1mg/kg, just above the detection threshold for brain cholinesterase inhibition. We assessed long-term effects from adolescence (PN30) through full adulthood (PN150), measuring the expression of serotonin receptors and serotonin turnover (index of presynaptic impulse activity) in cerebrocortical brain regions encompassing the projections that are known targets for nicotine and chlorpyrifos. Nicotine or chlorpyrifos individually increased the expression of serotonin receptors, with greater effects on males than on females and with distinct temporal and regional patterns indicative of adaptive synaptic changes rather than simply an extension of initial injury. This interpretation was confirmed by our finding an increase in serotonin turnover, connoting presynaptic serotonergic hyperactivity. Animals receiving the combined treatment showed a reduction in these adaptive effects on receptor binding and turnover relative to the individual agents, or even an effect in the opposite direction; further, normal sex differences in serotonin receptor concentrations were dissipated or reversed, an effect that was confirmed by behavioral evaluations in the Novel Objection Recognition Test. In addition to the known liabilities associated with maternal smoking during pregnancy, our results point to additional costs in the form of heightened vulnerability to neurotoxic chemicals encountered later in life. PMID:25592617

  19. ErbB polymorphisms: insights and implications for response to targeted cancer therapeutics.

    Science.gov (United States)

    Alaoui-Jamali, Moulay A; Morand, Grégoire B; da Silva, Sabrina Daniela

    2015-01-01

    Advances in high-throughput genomic-scanning have expanded the repertory of genetic variations in DNA sequences encoding ErbB tyrosine kinase receptors in humans, including single nucleotide polymorphisms (SNPs), polymorphic repetitive elements, microsatellite variations, small-scale insertions and deletions. The ErbB family members: EGFR, ErbB2, ErbB3, and ErbB4 receptors are established as drivers of many aspects of tumor initiation and progression to metastasis. This knowledge has provided rationales for the development of an arsenal of anti-ErbB therapeutics, ranging from small molecule kinase inhibitors to monoclonal antibodies. Anti-ErbB agents are becoming the cornerstone therapeutics for the management of cancers that overexpress hyperactive variants of ErbB receptors, in particular ErbB2-positive breast cancer and non-small cell lung carcinomas. However, their clinical benefit has been limited to a subset of patients due to a wide heterogeneity in drug response despite the expression of the ErbB targets, attributed to intrinsic (primary) and to acquired (secondary) resistance. Somatic mutations in ErbB tyrosine kinase domains have been extensively investigated in preclinical and clinical setting as determinants for either high sensitivity or resistance to anti-ErbB therapeutics. In contrast, only scant information is available on the impact of SNPs, which are widespread in genes encoding ErbB receptors, on receptor structure and activity, and their predictive values for drug susceptibility. This review aims to briefly update polymorphic variations in genes encoding ErbB receptors based on recent advances in deep sequencing technologies, and to address challenging issues for a better understanding of the functional impact of single versus combined SNPs in ErbB genes to receptor topology, receptor-drug interaction, and drug susceptibility. The potential of exploiting SNPs in the era of stratified targeted therapeutics is discussed. PMID:25699077

  20. ErbB polymorphisms: Insights and implications for response to targeted cancer therapeutics

    Directory of Open Access Journals (Sweden)

    Moulay A Alaoui-Jamali

    2015-02-01

    Full Text Available Advances in high-throughput genomic-scanning have expanded the repertory of genetic variations in DNA sequences encoding ErbB tyrosine kinase receptors in humans, including single nucleotide polymorphisms (SNPs, polymorphic repetitive elements, microsatellite variations, small-scale insertions and deletions. The ErbB family members: EGFR, ErbB2, ErbB3 and ErbB4 receptors are established as drivers of many aspects of tumor initiation and progression to metastasis. This knowledge has provided rationales for the development of an arsenal of anti-ErbB therapeutics, ranging from small molecule kinase inhibitors to monoclonal antibodies. Anti-ErbB agents are becoming the cornerstone therapeutics for the management of cancers that overexpress hyperactive variants of ErbB receptors, in particular ErbB2-positive breast cancer and non-small cell lung carcinomas. However, their clinical benefit has been limited to a subset of patients due to a wide heterogeneity in drug response despite the expression of the ErbB targets, attributed to intrinsic (primary and to acquired (secondary resistance. Somatic mutations in ErbB tyrosine kinase domains have been extensively investigated in preclinical and clinical setting as determinants for either high sensitivity or resistance to anti-ErbB therapeutics. In contrast, only scant information is available on the impact of SNPs, which are widespread in genes encoding ErbB receptors, on receptor structure and activity, and their predictive values for drug susceptibility. This review aims to briefly update polymorphic variations in genes encoding ErbB receptors based on recent advances in deep sequencing technologies, and to address challenging issues for a better understanding of the functional impact of single versus combined SNPs in ErbB genes to receptor topology, receptor-drug interaction, and drug susceptibility. The potential of exploiting SNPs in the era of stratified targeted therapeutics is discussed.

  1. The Response Regulator BfmR Is a Potential Drug Target for Acinetobacter baumannii.

    Science.gov (United States)

    Russo, Thomas A; Manohar, Akshay; Beanan, Janet M; Olson, Ruth; MacDonald, Ulrike; Graham, Jessica; Umland, Timothy C

    2016-01-01

    Identification and validation is the first phase of target-based antimicrobial development. BfmR (RstA), a response regulator in a two-component signal transduction system (TCS) in Acinetobacter baumannii, is an intriguing potential antimicrobial target. A unique characteristic of BfmR is that its inhibition would have the dual benefit of significantly decreasing in vivo survival and increasing sensitivity to selected antimicrobials. Studies on the clinically relevant strain AB307-0294 have shown BfmR to be essential in vivo. Here, we demonstrate that this phenotype in strains AB307-0294 and AB908 is mediated, in part, by enabling growth in human ascites fluid and serum. Further, BfmR conferred resistance to complement-mediated bactericidal activity that was independent of capsular polysaccharide. Importantly, BfmR also increased resistance to the clinically important antimicrobials meropenem and colistin. BfmR was highly conserved among A. baumannii strains. The crystal structure of the receiver domain of BfmR was determined, lending insight into putative ligand binding sites. This enabled an in silico ligand binding analysis and a blind docking strategy to assess use as a potential druggable target. Predicted binding hot spots exist at the homodimer interface and the phosphorylation site. These data support pursuing the next step in the development process, which includes determining the degree of inhibition needed to impact growth/survival and the development a BfmR activity assay amenable to high-throughput screening for the identification of inhibitors. Such agents would represent a new class of antimicrobials active against A. baumannii which could be active against other Gram-negative bacilli that possess a TCS with shared homology. IMPORTANCE Increasing antibiotic resistance in bacteria, particularly Gram-negative bacilli, has significantly affected the ability of physicians to treat infections, with resultant increased morbidity, mortality, and health

  2. Overview of affinity biosensors in food analysis.

    Science.gov (United States)

    Patel, Pradip D

    2006-01-01

    The 4 major driving forces that are expected to lead to increased use of affinity biosensors that meet crucial industrial test specifications, e.g., fast, reliable, cost-effective, and use of low-skilled personnel, are (1) strict legislative framework, e.g., recent changes proposed to the European food safety and hygiene legislation, EC No. 178/2002; (2) industrial shift from quality control to quality assurance procedures, e.g., Hazard Analysis Critical Control Point, ensuring effective positioning in the global competitive trade; (3) just-in-time production resulting in 'right' product every time; and (4) consumer demand for safe and wholesome products. The affinity biosensors field has expanded significantly over the past decade, with a projected global biosensors market growth from $6.1 billion in 2004 to $8.2 billion in 2009, representing major industrial sectors (e.g., Pharma, Medicare, and Food). This brief review is targeted to affinity biosensors developed for the food industry and includes research and development leading to biosensors for microbiological and chemical analytes of industrial concern, commercial biosensors products on the market, and examples of future prospects in this diagnostic field. PMID:16792079

  3. Overview of affinity biosensors in food analysis.

    Science.gov (United States)

    Patel, Pradip D

    2006-01-01

    The 4 major driving forces that are expected to lead to increased use of affinity biosensors that meet crucial industrial test specifications, e.g., fast, reliable, cost-effective, and use of low-skilled personnel, are (1) strict legislative framework, e.g., recent changes proposed to the European food safety and hygiene legislation, EC No. 178/2002; (2) industrial shift from quality control to quality assurance procedures, e.g., Hazard Analysis Critical Control Point, ensuring effective positioning in the global competitive trade; (3) just-in-time production resulting in 'right' product every time; and (4) consumer demand for safe and wholesome products. The affinity biosensors field has expanded significantly over the past decade, with a projected global biosensors market growth from $6.1 billion in 2004 to $8.2 billion in 2009, representing major industrial sectors (e.g., Pharma, Medicare, and Food). This brief review is targeted to affinity biosensors developed for the food industry and includes research and development leading to biosensors for microbiological and chemical analytes of industrial concern, commercial biosensors products on the market, and examples of future prospects in this diagnostic field.

  4. Realization of Fractal Affine Transformation

    Institute of Scientific and Technical Information of China (English)

    2001-01-01

    This paper gives the definition of fractal affine transformation and presents a specific method for its realization and its cor responding mathematical equations which are essential in fractal image construction.

  5. Representations of affine Hecke algebras

    CERN Document Server

    Xi, Nanhua

    1994-01-01

    Kazhdan and Lusztig classified the simple modules of an affine Hecke algebra Hq (q E C*) provided that q is not a root of 1 (Invent. Math. 1987). Ginzburg had some very interesting work on affine Hecke algebras. Combining these results simple Hq-modules can be classified provided that the order of q is not too small. These Lecture Notes of N. Xi show that the classification of simple Hq-modules is essentially different from general cases when q is a root of 1 of certain orders. In addition the based rings of affine Weyl groups are shown to be of interest in understanding irreducible representations of affine Hecke algebras. Basic knowledge of abstract algebra is enough to read one third of the book. Some knowledge of K-theory, algebraic group, and Kazhdan-Lusztig cell of Cexeter group is useful for the rest

  6. Infinite transitivity on affine varieties

    OpenAIRE

    Arzhantsev, Ivan; Flenner, Hubert; Kaliman, Shulim; Kutzschebauch, Frank; ZAIDENBERG, MIKHAIL

    2012-01-01

    In this note we survey recent results on automorphisms of affine algebraic varieties, infinitely transitive group actions and flexibility. We present related constructions and examples, and discuss geometric applications and open problems.

  7. Targeting brains, producing responsibilities: the use of neuroscience within British social policy.

    Science.gov (United States)

    Broer, Tineke; Pickersgill, Martyn

    2015-05-01

    Concepts and findings 'translated' from neuroscientific research are finding their way into UK health and social policy discourse. Critical scholars have begun to analyse how policies tend to 'misuse' the neurosciences and, further, how these discourses produce unwarranted and individualizing effects, rooted in middle-class values and inducing guilt and anxiety. In this article, we extend such work while simultaneously departing from the normative assumptions implied in the concept of 'misuse'. Through a documentary analysis of UK policy reports focused on the early years, adolescence and older adults, we examine how these employ neuroscientific concepts and consequently (re)define responsibility. In the documents analysed, responsibility was produced in three different but intersecting ways: through a focus on optimisation, self-governance, and vulnerability. Our work thereby adds to social scientific examinations of neuroscience in society that show how neurobiological terms and concepts can be used to construct and support a particular imaginary of citizenship and the role of the state. Neuroscience may be leveraged by policy makers in ways that (potentially) reduce the target of their intervention to the soma, but do so in order to expand the outcome of the intervention to include the enhancement of society writ large. By attending as well to more critical engagements with neuroscience in policy documents, our analysis demonstrates the importance of being mindful of the limits to the deployment of a neurobiological idiom within policy settings. Accordingly, we contribute to increased empirical specificity concerning the impacts and translation of neuroscientific knowledge in contemporary society whilst refusing to take for granted the idea that the neurosciences necessarily have a dominant role (to play). PMID:25792340

  8. NBR1-mediated selective autophagy targets insoluble ubiquitinated protein aggregates in plant stress responses.

    Directory of Open Access Journals (Sweden)

    Jie Zhou

    Full Text Available Plant autophagy plays an important role in delaying senescence, nutrient recycling, and stress responses. Functional analysis of plant autophagy has almost exclusively focused on the proteins required for the core process of autophagosome assembly, but little is known about the proteins involved in other important processes of autophagy, including autophagy cargo recognition and sequestration. In this study, we report functional genetic analysis of Arabidopsis NBR1, a homolog of mammalian autophagy cargo adaptors P62 and NBR1. We isolated two nbr1 knockout mutants and discovered that they displayed some but not all of the phenotypes of autophagy-deficient atg5 and atg7 mutants. Like ATG5 and ATG7, NBR1 is important for plant tolerance to heat, oxidative, salt, and drought stresses. The role of NBR1 in plant tolerance to these abiotic stresses is dependent on its interaction with ATG8. Unlike ATG5 and ATG7, however, NBR1 is dispensable in age- and darkness-induced senescence and in resistance to a necrotrophic pathogen. A selective role of NBR1 in plant responses to specific abiotic stresses suggest that plant autophagy in diverse biological processes operates through multiple cargo recognition and delivery systems. The compromised heat tolerance of atg5, atg7, and nbr1 mutants was associated with increased accumulation of insoluble, detergent-resistant proteins that were highly ubiquitinated under heat stress. NBR1, which contains an ubiquitin-binding domain, also accumulated to high levels with an increasing enrichment in the insoluble protein fraction in the autophagy-deficient mutants under heat stress. These results suggest that NBR1-mediated autophagy targets ubiquitinated protein aggregates most likely derived from denatured or otherwise damaged nonnative proteins generated under stress conditions.

  9. pH-responsive hybrid quantum dots for targeting hypoxic tumor siRNA delivery.

    Science.gov (United States)

    Zhu, HongYan; Zhang, ShengYu; Ling, Yong; Meng, GuoLiang; Yang, Yu; Zhang, Wei

    2015-12-28

    Hypoxia is a characteristic of cancer and plays a key role in tumorigenesis, angiogenesis and resistance to cancer therapies. SiRNA treatment is effective against hypoxic tumors by gene silencing. However, siRNA delivery to the hypoxic regions of solid tumors still presents a challenge due to the distance from blood vessels and the increased presence of efflux transporters. Therefore, tumor therapies would be improved through the immediate development of an effective siRNA delivery system to hypoxic regions. To this end, we synthesized a system to deliver HIF-1α siRNA into hypoxic tumor cells. The system consists of a functional shell composed of 2-deoxyglucose (DG)-polyethylene glycol (PEG) connected with the compound of lipoic acid, lysine and 9-poly-d-arginine (LA-Lys-9R) by a hydrazone bond and a core of CdTe quantum dots (QDs). The molecular structure of DG-PEG-LA-Lys-9R was confirmed by liquid chromatography-mass spectrometry (LC-MS), nuclear magnetic resonance (NMR) spectroscopy, Fourier transform infrared spectroscopy (FTIR), and sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). The multifunctional CdTe QDs measured approximately 200 nm and showed excellent biocompatibility, perfect siRNA binding capability and enhanced hypoxic tumor targeting. Importantly, the system described here is pH-responsive with a hydrazone bond; therefore, it avoids GLUT1 receptor-mediated endocytic recycling, resulting in irreversible delivery of the siRNA. We used Western blots to confirm the superior gene silencing efficiency induced by the DG-PEG-LA-Lys-9R with hydrazone modified CdTe QDs. Here, we demonstrate high efficacy of the siRNA tumor delivery system using in vitro and in vivo experiments. In addition, these studies demonstrate that pH-responsive hybrid quantum dots show improved antitumor efficacy with decreased organ toxicity, indicating a promising siRNA delivery system for hypoxic cancer therapy. PMID:26590349

  10. Inflammatory and immune responses in the cochlea: potential therapeutic targets for sensorineural hearing loss

    Directory of Open Access Journals (Sweden)

    Masato eFujioka

    2014-12-01

    Full Text Available The inner ear was previously assumed to be an immune-privileged organ due to the existence of its tight junction-based blood-labyrinth barrier. However, studies performed during the past decade revealed that the mesenchymal region of the cochlea, including its lateral wall, is a common site of inflammation. Neutrophils do not enter this region, which is consistent with the old dogma; however, bone marrow-derived resident macrophages are always present in the spiral ligament of the lateral wall and are activated in response to various types of insults, including noise exposure, ischemia, mitochondrial damage and surgical stress. Recent studies have also revealed another type of immune cell, called perivascular melanocyte-like macrophages (PVM/Ms, in the stria vascularis. These dedicated antigen-presenting cells also control vascular contraction and permeability. This review discusses a series of reports regarding inflammatory/immune cells in the cochlear lateral wall, the pathways involved in cochlear damage and their potential as therapeutic targets.

  11. Identification of drought-responsive compounds in potato through a combined transcriptomic and targeted metabolite approach.

    Science.gov (United States)

    Evers, Danièle; Lefèvre, Isabelle; Legay, Sylvain; Lamoureux, Didier; Hausman, Jean-François; Rosales, Raymundo Oscar Gutierrez; Marca, Luz Rosalina Tincopa; Hoffmann, Lucien; Bonierbale, Merideth; Schafleitner, Roland

    2010-05-01

    Two potato clones (Solanum tuberosum L.) of the Andean cultivar group, called Sullu and SS2613, with different drought-tolerance phenotypes were exposed to a continuously increasing drought stress in a field trial. At the physiological level, while relative leaf water contents were similar in both clones, osmotic potential was lower in Sullu and declined more strongly during drought compared with SS2613. In the drought-stressed plants, tuber yield was reduced by about 70% compared with control plants in both clones. Potato cDNA microarrays and target metabolite analysis were performed on leaves sampled at several time-points after the onset of drought. At the transcriptomic level, photosynthesis-related genes were already strongly repressed in Sullu after 28 d of withholding irrigation and even more strongly after a longer stress duration, whereas, in SS2613, repression occurred only after 49 d of soil drying; similarly, a strong perturbation of carbohydrate-related genes was observed in Sullu. At the metabolite level, differential accumulation of osmotically active solutes was observed between the two cultivars; indeed, in Sullu, contents of galactose, inositol, galactinol, proline, and proline analogues were higher upon drought stress compared with SS2613. These results point to different drought responses in the cultivars at the leaf level, with, however, similar tuber yield reductions. The previously shown tolerant clone Sullu lost part of its tolerance under the experimental conditions used here; it was, however, able to maintain an absolute yield three times higher than SS2613.

  12. A thermodynamic approach to the affinity optimization of drug candidates.

    Science.gov (United States)

    Freire, Ernesto

    2009-11-01

    High throughput screening and other techniques commonly used to identify lead candidates for drug development usually yield compounds with binding affinities to their intended targets in the mid-micromolar range. The affinity of these molecules needs to be improved by several orders of magnitude before they become viable drug candidates. Traditionally, this task has been accomplished by establishing structure activity relationships to guide chemical modifications and improve the binding affinity of the compounds. As the binding affinity is a function of two quantities, the binding enthalpy and the binding entropy, it is evident that a more efficient optimization would be accomplished if both quantities were considered and improved simultaneously. Here, an optimization algorithm based upon enthalpic and entropic information generated by Isothermal Titration Calorimetry is presented.

  13. Rationally designed inhibitor targeting antigen-trimming aminopeptidases enhances antigen presentation and cytotoxic T-cell responses.

    Science.gov (United States)

    Zervoudi, Efthalia; Saridakis, Emmanuel; Birtley, James R; Seregin, Sergey S; Reeves, Emma; Kokkala, Paraskevi; Aldhamen, Yasser A; Amalfitano, Andrea; Mavridis, Irene M; James, Edward; Georgiadis, Dimitris; Stratikos, Efstratios

    2013-12-01

    Intracellular aminopeptidases endoplasmic reticulum aminopeptidases 1 and 2 (ERAP1 and ERAP2), and as well as insulin-regulated aminopeptidase (IRAP) process antigenic epitope precursors for loading onto MHC class I molecules and regulate the adaptive immune response. Their activity greatly affects the antigenic peptide repertoire presented to cytotoxic T lymphocytes and as a result can regulate cytotoxic cellular responses contributing to autoimmunity or immune evasion by viruses and cancer cells. Therefore, pharmacological regulation of their activity is a promising avenue for modulating the adaptive immune response with possible applications in controlling autoimmunity, in boosting immune responses to pathogens, and in cancer immunotherapy. In this study we exploited recent structural and biochemical analysis of ERAP1 and ERAP2 to design and develop phosphinic pseudopeptide transition state analogs that can inhibit this family of enzymes with nM affinity. X-ray crystallographic analysis of one such inhibitor in complex with ERAP2 validated our design, revealing a canonical mode of binding in the active site of the enzyme, and highlighted the importance of the S2' pocket for achieving inhibitor potency. Antigen processing and presentation assays in HeLa and murine colon carcinoma (CT26) cells showed that these inhibitors induce increased cell-surface antigen presentation of transfected and endogenous antigens and enhance cytotoxic T-cell responses, indicating that these enzymes primarily destroy epitopes in those systems. This class of inhibitors constitutes a promising tool for controlling the cellular adaptive immune response in humans by modulating the antigen processing and presentation pathway. PMID:24248368

  14. Measurement Tools and Target Symptoms/Skills Used to Assess Treatment Response for Individuals with Autism Spectrum Disorder

    Science.gov (United States)

    Bolte, Erin Elizabeth; Diehl, Joshua John

    2013-01-01

    This study examined the measurement tools and target symptoms/skills used to assess treatment response during Autism Spectrum Disorder (ASD) intervention trials from 2001 through 2010. Data from 195 prospective trials were analyzed. There were 289 unique measurement tools, of which 61.6% were used only once, and 20.8 % were investigator-designed.…

  15. PET Imaging in Head and Neck Cancer Patients to Monitor Treatment Response: A Future Role for EGFR-Targeted Imaging

    NARCIS (Netherlands)

    Dijk, L.K. van; Boerman, O.C.; Kaanders, J.H.A.M.; Bussink, J.

    2015-01-01

    Approximately 50,000 new cases of head and neck squamous cell carcinoma (HNSCC) are diagnosed worldwide each year and subsequently treated with surgery, chemotherapy, radiotherapy, and/or targeted therapy. The heterogeneity of the patient population in terms of treatment response drives the search f

  16. Glutathione-responsive nano-vehicles as a promising platform for targeted intracellular drug and gene delivery

    NARCIS (Netherlands)

    Cheng, Ru; Feng, Fang; Meng, Fenghua; Deng, Chao; Feijen, Jan; Zhong, Zhiyuan

    2011-01-01

    The past couple of years have witnessed a tremendous progress in the development of glutathione-responsive nano-vehicles for targeted intracellular drug and gene delivery, as driven by the facts that (i) many therapeutics (e.g. anti-cancer drugs, photosensitizers, and anti-oxidants) and biotherapeut

  17. A Comprehensive Analysis of the Dynamic Response to Aphidicolin-Mediated Replication Stress Uncovers Targets for ATM and ATMIN

    Directory of Open Access Journals (Sweden)

    Abdelghani Mazouzi

    2016-04-01

    Full Text Available The cellular response to replication stress requires the DNA-damage-responsive kinase ATM and its cofactor ATMIN; however, the roles of this signaling pathway following replication stress are unclear. To identify the functions of ATM and ATMIN in response to replication stress, we utilized both transcriptomics and quantitative mass-spectrometry-based phosphoproteomics. We found that replication stress induced by aphidicolin triggered widespread changes in both gene expression and protein phosphorylation patterns. These changes gave rise to distinct early and late replication stress responses. Furthermore, our analysis revealed previously unknown targets of ATM and ATMIN downstream of replication stress. We demonstrate ATMIN-dependent phosphorylation of H2AX and of CRMP2, a protein previously implicated in Alzheimer’s disease but not in the DNA damage response. Overall, our dataset provides a comprehensive resource for discovering the cellular responses to replication stress and, potentially, associated pathologies.

  18. Explaining the effects of targeted online advertising on children's cognitive, affective, and behavioral brand responses

    NARCIS (Netherlands)

    E. van Reijmersdal; E. Rozendaal; N. Smink; G. van Noort; M. Buijzen

    2013-01-01

    Increasingly, information from children's profile pages on social network sites is used to target online advertising. This practice has raised concerns in society and academia, however, effects of profile targeting on children remained unstudied. Therefore, this study focused on children's cognitive

  19. Quiet Eye Duration Is Responsive to Variability of Practice and to the Axis of Target Changes

    Science.gov (United States)

    Horn, Robert R.; Okumura, Michelle S.; Alexander, Melissa G. F.; Gardin, Fredrick A.; Sylvester, Curtis T.

    2012-01-01

    We tested the hypothesis that quiet eye, the final fixation before the initiation of a movement in aiming tasks, is used to scale the movement's parameters. Two groups of 12 participants (N = 24) threw darts to targets in the horizontal and vertical axes under conditions of higher (random) or lower (blocked) target variability. Supporting our…

  20. The classical origin of quantum affine algebra in squashed sigma models

    OpenAIRE

    Kawaguchi, Io; Matsumoto, Takuya; Yoshida, Kentaroh

    2012-01-01

    We consider a quantum affine algebra realized in two-dimensional non-linear sigma models with target space three-dimensional squashed sphere. Its affine generators are explicitly constructed and the Poisson brackets are computed. The defining relations of quantum affine algebra in the sense of the Drinfeld first realization are satisfied at classical level. The relation to the Drinfeld second realization is also discussed including higher conserved charges. Finally we comment on a semiclassic...

  1. Time-Frequency Analysis of Electromagnetic Pulse Response from a Spherical Target

    Institute of Scientific and Technical Information of China (English)

    陈东; 金亚秋

    2003-01-01

    Transient backscattering from a spherical target under incidence of an electromagnetic short pulse is studied. The target can be a perfectly conducting sphere, a dielectric sphere or a dielectric spherical shell. To understand the scattering mechanism from transient impulse echoes for target detection, both the short-time Fourier transform(STFT) and the wavelet transform (WT) are applied to retrieval of scattering information from the backscattering data. Analysis in both the time and frequency domains demonstrates that the WT is more feasible than the STFT to clarifying scattering process of the scatterer because of its excellent multi-resolution characteristic. This technique shall be helpful for scattering analysis and detection of more complex single or multi-targets.

  2. Differential Inflammatory Response to Inhaled Lipopolysaccharide Targeted Either to the Airways or the Alveoli in Man

    OpenAIRE

    Möller, Winfried; Heimbeck, Irene; Hofer, Thomas P J; Khadem Saba, Gülnaz; Neiswirth, Margot; Frankenberger, Marion; Ziegler-Heitbrock, Löms

    2012-01-01

    Endotoxin (Lipopolysaccharide, LPS) is a potent inducer of inflammation and there is various LPS contamination in the environment, being a trigger of lung diseases and exacerbation. The objective of this study was to assess the time course of inflammation and the sensitivities of the airways and alveoli to targeted LPS inhalation in order to understand the role of LPS challenge in airway disease. In healthy volunteers without any bronchial hyperresponsiveness we targeted sequentially 1, 5 and...

  3. The Salmonella Effector Protein SopA Modulates Innate Immune Responses by Targeting TRIM E3 Ligase Family Members.

    Directory of Open Access Journals (Sweden)

    Jana Kamanova

    2016-04-01

    Full Text Available Salmonella Typhimurium stimulates inflammatory responses in the intestinal epithelium, which are essential for its ability to replicate within the intestinal tract. Stimulation of these responses is strictly dependent on the activity of a type III secretion system encoded within its pathogenicity island 1, which through the delivery of effector proteins, triggers signaling pathways leading to inflammation. One of these effectors is SopA, a HECT-type E3 ligase, which is required for the efficient stimulation of inflammation in an animal model of Salmonella Typhimurium infection. We show here that SopA contributes to the stimulation of innate immune responses by targeting two host E3 ubiquitin ligases, TRIM56 and TRIM65. We also found that TRIM65 interacts with the innate immune receptor MDA5 enhancing its ability to stimulate interferon-β signaling. Therefore, by targeting TRIM56 and TRIM65, SopA can stimulate signaling through two innate immune receptors, RIG-I and MDA5. These findings describe a Salmonella mechanism to modulate inflammatory responses by directly targeting innate immune signaling mechanisms.

  4. Involvement of miR160/miR393 and their targets in cassava responses to anthracnose disease.

    Science.gov (United States)

    Pinweha, Nattaya; Asvarak, Thipa; Viboonjun, Unchera; Narangajavana, Jarunya

    2015-02-01

    Cassava is a starchy root crop for food and industrial applications in many countries around the world. Among the factors that affect cassava production, diseases remain the major cause of yield loss. Cassava anthracnose disease is caused by the fungus Colletotrichum gloeosporioides. Severe anthracnose attacks can cause tip die-backs and stem cankers, which can affect the availability of planting materials especially in large-scale production systems. Recent studies indicate that plants over- or under-express certain microRNAs (miRNAs) to cope with various stresses. Understanding how a disease-resistant plant protects itself from pathogens should help to uncover the role of miRNAs in the plant immune system. In this study, the disease severity assay revealed different response to C. gloeosporioides infection in two cassava cultivars. Quantitative RT-PCR analysis uncovered the differential expression of the two miRNAs and their target genes in the two cassava cultivars that were subjected to fungal infection. The more resistant cultivar revealed the up-regulation of miR160 and miR393, and consequently led to low transcript levels in their targets, ARF10 and TIR1, respectively. The more susceptible cultivar exhibited the opposite pattern. The cis-regulatory elements relevant to defense and stress responsiveness, fungal elicitor responsiveness and hormonal responses were the most prevalent present in the miRNAs gene promoter regions. The possible dual role of these specific miRNAs and their target genes associated with cassava responses to C. gloeosporioides is discussed. This is the first study to address the molecular events by which miRNAs which might play a role in fungal-infected cassava. A better understanding of the functions of miRNAs target genes should greatly increase our knowledge of the mechanism underlying susceptibility and lead to new strategies to enhance disease tolerance in this economically important crop.

  5. Genome-wide identification of Brassica napus microRNAs and their targets in response to cadmium.

    Science.gov (United States)

    Zhou, Zhao Sheng; Song, Jian Bo; Yang, Zhi Min

    2012-07-01

    MicroRNAs (miRNAs) are a distinct class of small RNAs in plants that not only regulate biological processes but also regulate response to environmental stresses. The toxic heavy metal cadmium (Cd) induces expression of several miRNAs in rapeseed (Brassica napus), but it is not known on a genome-wide scale how the expression of miRNAs and their target genes, is regulated by Cd. In this study, four small RNA libraries and four degradome libraries were constructed from Cd-treated and non-Cd-treated roots and shoots of B. napus seedlings. Using high-throughput sequencing, the study identified 84 conserved and non-conserved miRNAs (belonging to 37 miRNA families) from Cd-treated and non-treated B. napus, including 19 miRNA members that were not identified before. Some of the miRNAs were validated by RNA gel blotting. Most of the identified miRNAs were found to be differentially expressed in roots/shoots or regulated by Cd exposure. The study simultaneously identified 802 targets for the 37 (24 conserved and 13 non-conserved) miRNA families, from which there are 200, 537, and 65 targets, belonging to categories I, II, and III, respectively. In category I alone, many novel targets for miRNAs were identified and shown to be involved in plant response to Cd.

  6. Proteome-wide analysis of SUMO2 targets in response to pathological DNA replication stress in human cells

    DEFF Research Database (Denmark)

    Bursomanno, Sara; Beli, Petra; Khan, Asif M;

    2015-01-01

    subfamily. SUMO2/3, in contrast to SUMO1, are predominantly involved in the cellular response to certain stresses, including heat shock. Substantial evidence from studies in yeast has shown that SUMOylation plays an important role in the regulation of DNA replication and repair. Here, we report a proteomic...... analysis of proteins modified by SUMO2 in response to DNA replication stress in S phase in human cells. We have identified a panel of 22 SUMO2 targets with increased SUMOylation during DNA replication stress, many of which play key functions within the DNA replication machinery and/or in the cellular...

  7. Analysis of cellular responses of macrophages to zinc ions and zinc oxide nanoparticles: a combined targeted and proteomic approach

    Science.gov (United States)

    Triboulet, Sarah; Aude-Garcia, Catherine; Armand, Lucie; Gerdil, Adèle; Diemer, Hélène; Proamer, Fabienne; Collin-Faure, Véronique; Habert, Aurélie; Strub, Jean-Marc; Hanau, Daniel; Herlin, Nathalie; Carrière, Marie; van Dorsselaer, Alain; Rabilloud, Thierry

    2014-05-01

    Two different zinc oxide nanoparticles, as well as zinc ions, are used to study the cellular responses of the RAW 264 macrophage cell line. A proteomic screen is used to provide a wide view of the molecular effects of zinc, and the most prominent results are cross-validated by targeted studies. Furthermore, the alteration of important macrophage functions (e.g. phagocytosis) by zinc is also investigated. The intracellular dissolution/uptake of zinc is also studied to further characterize zinc toxicity. Zinc oxide nanoparticles dissolve readily in the cells, leading to high intracellular zinc concentrations, mostly as protein-bound zinc. The proteomic screen reveals a rather weak response in the oxidative stress response pathway, but a strong response both in the central metabolism and in the proteasomal protein degradation pathway. Targeted experiments confirm that carbohydrate catabolism and proteasome are critical determinants of sensitivity to zinc, which also induces DNA damage. Conversely, glutathione levels and phagocytosis appear unaffected at moderately toxic zinc concentrations.Two different zinc oxide nanoparticles, as well as zinc ions, are used to study the cellular responses of the RAW 264 macrophage cell line. A proteomic screen is used to provide a wide view of the molecular effects of zinc, and the most prominent results are cross-validated by targeted studies. Furthermore, the alteration of important macrophage functions (e.g. phagocytosis) by zinc is also investigated. The intracellular dissolution/uptake of zinc is also studied to further characterize zinc toxicity. Zinc oxide nanoparticles dissolve readily in the cells, leading to high intracellular zinc concentrations, mostly as protein-bound zinc. The proteomic screen reveals a rather weak response in the oxidative stress response pathway, but a strong response both in the central metabolism and in the proteasomal protein degradation pathway. Targeted experiments confirm that carbohydrate

  8. A role for plasma cell targeting agents in immune tolerance induction in autoimmune disease and antibody responses to therapeutic proteins.

    Science.gov (United States)

    Rosenberg, A S; Pariser, A R; Diamond, B; Yao, L; Turka, L A; Lacana, E; Kishnani, P S

    2016-04-01

    Antibody responses to life saving therapeutic protein products, such as enzyme replacement therapies (ERT) in the setting of lysosomal storage diseases, have nullified product efficacy and caused clinical deterioration and death despite treatment with immune-suppressive therapies. Moreover, in some autoimmune diseases, pathology is mediated by a robust antibody response to endogenous proteins such as is the case in pulmonary alveolar proteinosis, mediated by antibodies to Granulocyte Macrophage-Colony Stimulating Factor (GM-CSF). In this work, we make the case that in such settings, when the antibody response is high titered, sustained, and refractory to immune suppressive treatments, the antibody response is mediated by long-lived plasma cells which are relatively unperturbed by immune suppressants including rituximab. However, long-lived plasma cells can be targeted by proteasome inhibitors such as bortezomib. Recent reports of successful reversal of antibody responses with bortezomib in the settings of ERT and Thrombotic Thrombocytopenic Purpura (TTP) argue that the safety and efficacy of such plasma cell targeting agents should be evaluated in larger scale clinical trials to delineate the risks and benefits of such therapies in the settings of antibody-mediated adverse effects to therapeutic proteins and autoantibody mediated pathology. PMID:26928739

  9. TARGET COSTING – THE RESPONSE OF THE MANAGERIAL ACCOUNTING TO CHANGES IN THE ENVIRONMENT

    Directory of Open Access Journals (Sweden)

    OFILEANU DIMI

    2014-06-01

    Full Text Available The aim of this article is to highlight the Target Costing concept and the way in which the concept is used by the Romanian entities. The article analyzes the concept starting from the time of its appearance and up to the present moment, presenting the concept’s characteristics and its mode of action. A careful review of the specialized literature will be presented and the fact that this concept represents in fact a method of management. In order to study an eventual correlation between using Target Costing concept and the financial results obtained, an analysis of a sample of Romanian entities which produce shoes will be made. Also, it was analyzed the level of use and how much the Target Costing concept is known.

  10. Assessment of targeted and non-targeted responses in cells deficient in ATM function following exposure to low and high dose X-rays.

    Science.gov (United States)

    Kiuru, Anne; Kämäräinen, Meerit; Heinävaara, Sirpa; Pylkäs, Katri; Chapman, Kim; Koivistoinen, Armi; Parviainen, Teuvo; Winqvist, Robert; Kadhim, Munira; Launonen, Virpi; Lindholm, Carita

    2014-01-01

    Radiation sensitivity at low and high dose exposure to X-rays was investigated by means of chromosomal aberration (CA) analysis in heterozygous ATM mutation carrier and A-T patient (biallelic ATM mutation) lymphoblastoid cell lines (LCLs). Targeted and non-targeted responses to acutely delivered irradiation were examined by applying a co-culture system that enables study of both directly irradiated cells and medium-mediated bystander effects in the same experimental setting. No indication of radiation hypersensitivity was observed at doses of 0.01 Gy or 0.1 Gy for the ATM mutation carrier LCL. The A-T patient cells also did not show low-dose response. There was significant increase in unstable CA yields for both ATM mutation carrier and A-T LCLs at 1 and 2 Gy, the A-T cells displaying more distinct dose dependency. Both chromosome and chromatid type aberrations were induced at an increased rate in the irradiated A-T cells, whereas for ATM carrier cells, only unstable chromosomal aberrations were increased above the level observed in the wild type cell line. No bystander effect could be demonstrated in any of the cell lines or doses applied. Characteristics typical for the A-T cell line were detected, i.e., high baseline frequency of CA that increased with dose. In addition, dose-dependent loss of cell viability was observed. In conclusion, CA analysis did not demonstrate low-dose (≤100 mGy) radiosensitivity in ATM mutation carrier cells or A-T patient cells. However, both cell lines showed increased radiosensitivity at high dose exposure.

  11. Assessment of targeted and non-targeted responses in cells deficient in ATM function following exposure to low and high dose X-rays.

    Directory of Open Access Journals (Sweden)

    Anne Kiuru

    Full Text Available Radiation sensitivity at low and high dose exposure to X-rays was investigated by means of chromosomal aberration (CA analysis in heterozygous ATM mutation carrier and A-T patient (biallelic ATM mutation lymphoblastoid cell lines (LCLs. Targeted and non-targeted responses to acutely delivered irradiation were examined by applying a co-culture system that enables study of both directly irradiated cells and medium-mediated bystander effects in the same experimental setting. No indication of radiation hypersensitivity was observed at doses of 0.01 Gy or 0.1 Gy for the ATM mutation carrier LCL. The A-T patient cells also did not show low-dose response. There was significant increase in unstable CA yields for both ATM mutation carrier and A-T LCLs at 1 and 2 Gy, the A-T cells displaying more distinct dose dependency. Both chromosome and chromatid type aberrations were induced at an increased rate in the irradiated A-T cells, whereas for ATM carrier cells, only unstable chromosomal aberrations were increased above the level observed in the wild type cell line. No bystander effect could be demonstrated in any of the cell lines or doses applied. Characteristics typical for the A-T cell line were detected, i.e., high baseline frequency of CA that increased with dose. In addition, dose-dependent loss of cell viability was observed. In conclusion, CA analysis did not demonstrate low-dose (≤100 mGy radiosensitivity in ATM mutation carrier cells or A-T patient cells. However, both cell lines showed increased radiosensitivity at high dose exposure.

  12. Molecular targets of omega 3 and conjugated linoleic fatty acids – micromanaging cellular response

    Directory of Open Access Journals (Sweden)

    Francesco eVisioli

    2012-02-01

    Full Text Available Essential fatty acids cannot be synthesized de novo by mammals and need to be ingested either with the diet or through the use of supplements/functional foods to ameliorate cardiovascular prognosis. This review focus on the molecular targets of omega 3 fatty acids and CLA, as paradigmatic molecules that can be explored both as nutrients and as pharmacological agents, especially as related to cardioprotection. In addition, we indicate novel molecular targets, namely microRNAs that might contribute to the observed biological activities of such essential fatty acids.

  13. Efficient mRNA-Based Genetic Engineering of Human NK Cells with High-Affinity CD16 and CCR7 Augments Rituximab-Induced ADCC against Lymphoma and Targets NK Cell Migration toward the Lymph Node-Associated Chemokine CCL19.

    Science.gov (United States)

    Carlsten, Mattias; Levy, Emily; Karambelkar, Amrita; Li, Linhong; Reger, Robert; Berg, Maria; Peshwa, Madhusudan V; Childs, Richard W

    2016-01-01

    For more than a decade, investigators have pursued methods to genetically engineer natural killer (NK) cells for use in clinical therapy against cancer. Despite considerable advances in viral transduction of hematopoietic stem cells and T cells, transduction efficiencies for NK cells have remained disappointingly low. Here, we show that NK cells can be genetically reprogramed efficiently using a cGMP-compliant mRNA electroporation method that induces rapid and reproducible transgene expression in nearly all transfected cells, without negatively influencing their viability, phenotype, and cytotoxic function. To study its potential therapeutic application, we used this approach to improve key aspects involved in efficient lymphoma targeting by adoptively infused ex vivo-expanded NK cells. Electroporation of NK cells with mRNA coding for the chemokine receptor CCR7 significantly promoted migration toward the lymph node-associated chemokine CCL19. Further, introduction of mRNA coding for the high-affinity antibody-binding receptor CD16 (CD16-158V) substantially augmented NK cell cytotoxicity against rituximab-coated lymphoma cells. Based on these data, we conclude that this approach can be utilized to genetically modify multiple modalities of NK cells in a highly efficient manner with the potential to improve multiple facets of their in vivo tumor targeting, thus, opening a new arena for the development of more efficacious adoptive NK cell-based cancer immunotherapies.

  14. Quantotypic Properties of QconCAT Peptides Targeting Bovine Host Response to Streptococcus uberis

    DEFF Research Database (Denmark)

    Bislev, Stine Lønnerup; Kusebauch, Ulrike; Codrea, Marius Cosmin;

    2012-01-01

    peptides from each of these 20 proteins were obtained through the QconCAT method. We present the quantotypic properties of these 40 proteotypic peptides, and discuss their application to research in host pathogen interactions. Our results clearly demonstrate a robust monitoring of 17 targeted host...

  15. Comparing Path Dependence and Spatial Targeting of Land Use in Implementing Climate Change Responses

    Directory of Open Access Journals (Sweden)

    Iain Brown

    2014-07-01

    Full Text Available Land use patterns are the consequence of dynamic processes that often include important legacy issues. Evaluation of past trends can be used to investigate the role of path dependence in influencing future land use through a reference “business as usual” (BAU scenario. These issues are explored with regard to objectives for woodland expansion in Scotland as a major pillar of climate change policy. Land use changes based upon recent trends and future transient scenarios to 2050 are used to assess viability of targets for reducing greenhouse gas emissions using analysis based on net emission change factors. The BAU scenario is compared with alternative future scenarios incorporating policy targets and stronger spatial targeting of land use change. Analysis highlights recent trends in new woodland planting on lower quality agricultural land due to socioeconomic and cultural factors. This land is mainly in the wetter uplands and often on carbon-rich soils. Woodland planting following this path dependence can therefore result in net carbon emissions for many years into the future due to soil disturbance during establishment. In contrast, alternative scenarios with more lowland woodland planting have net sequestration potential, with greatest benefits when carbon-rich soils are excluded from afforestation. Spatial targeting can also enhance other co-benefits such as habitat networks and climate change adaptation.

  16. Identification of drought-responsive microRNAs and their targets in Ammopiptanthus mongolicus by using high-throughput sequencing

    Science.gov (United States)

    Gao, Fei; Wang, Ning; Li, Huayun; Liu, Jisheng; Fu, Chenxi; Xiao, Zihua; Wei, Chunxiang; Lu, Xiaoduo; Feng, Jinchao; Zhou, Yijun

    2016-01-01

    MicroRNAs (miRNAs) regulate target gene expression to modulate plant growth, development, and biotic and abiotic stress response at the post-transcriptional level. Ammopiptanthus mongolicus, an ecologically important desert plant, is increasingly used as a model for studying stress tolerance in plants. The miRNA-mediated gene regulatory network might remarkably contribute to the high stress tolerance of A. mongolicus. However, a genome-wide identification of miRNAs and their targets is still lacking in A. mongolicus. In this study, 170 conserved and 156 non-conserved miRNAs were identified in A. mongolicus. We experimentally identified 298 miRNA-target pairs from the degradome data. Quantitative real-time polymerase chain reaction analyses identified 28 drought-responsive miRNAs in leaves and 15 in roots. Some characteristics of the miRNA-mediated regulatory network were found in A. mongolicus. Multiple miRNAs, including 2 newly identified non-conserved miRNAs, miR-P11 and miR-P14, generated from the precursors of miR169, were found to be involved in drought stress response. Further, miR2118 and miR858 participated in drought stress response by up-regulating OZF1 gene and certain MYB genes that were involved in the regulation of flavonol biosynthesis in A. mongolicus. The findings of this study might provide new insights for understanding the functions of miRNA in stress response in plants. PMID:27698373

  17. Affine density in wavelet analysis

    CERN Document Server

    Kutyniok, Gitta

    2007-01-01

    In wavelet analysis, irregular wavelet frames have recently come to the forefront of current research due to questions concerning the robustness and stability of wavelet algorithms. A major difficulty in the study of these systems is the highly sensitive interplay between geometric properties of a sequence of time-scale indices and frame properties of the associated wavelet systems. This volume provides the first thorough and comprehensive treatment of irregular wavelet frames by introducing and employing a new notion of affine density as a highly effective tool for examining the geometry of sequences of time-scale indices. Many of the results are new and published for the first time. Topics include: qualitative and quantitative density conditions for existence of irregular wavelet frames, non-existence of irregular co-affine frames, the Nyquist phenomenon for wavelet systems, and approximation properties of irregular wavelet frames.

  18. Properties of the Affine Invariant Ensemble Sampler in high dimensions

    CERN Document Server

    Huijser, David; Brewer, Brendon J

    2015-01-01

    We present theoretical and practical properties of the affine-invariant ensemble sampler Markov chain Monte Carlo method. In high dimensions the affine-invariant ensemble sampler shows unusual and undesirable properties. We demonstrate this with an $n$-dimensional correlated Gaussian toy problem with a known mean and covariance structure, and analyse the burn-in period. The burn-in period seems to be short, however upon closer inspection we discover the mean and the variance of the target distribution do not match the expected, known values. This problem becomes greater as $n$ increases. We therefore conclude that the affine-invariant ensemble sampler should be used with caution in high dimensional problems. We also present some theoretical results explaining this behaviour.

  19. Inhomogeneous self-affine carpets

    OpenAIRE

    Fraser, Jonathan M.

    2013-01-01

    We investigate the dimension theory of inhomogeneous self-affine carpets. Through the work of Olsen, Snigireva and Fraser, the dimension theory of inhomogeneous self-similar sets is now relatively well-understood, however, almost no progress has been made concerning more general non-conformal inhomogeneous attractors. If a dimension is countably stable, then the results are immediate and so we focus on the upper and lower box dimensions and compute these explicitly for large classes of inhomo...

  20. Protein isolation using affinity chromatography

    OpenAIRE

    Besselink, T.

    2012-01-01

    Many product or even waste streams in the food industry contain components that may have potential for e.g. functional foods. These streams are typically large in volume and the components of interest are only present at low concentrations. A robust and highly selective separation process should be developed for efficient isolation of the components. Affinity chromatography is such a selective method. Ligands immobilized to a stationary phase (e.g., a resin or membrane) are used to bind the c...

  1. Lectin affinity chromatography of glycolipids

    Energy Technology Data Exchange (ETDEWEB)

    Torres, B.V.; Smith, D.F.

    1987-05-01

    Since glycolipids (GLs) are either insoluble or form mixed micelles in water, lectin affinity chromatography in aqueous systems has not been applied to their separation. They have overcome this problem by using tetrahydrofuran (THF) in the mobile phase during chromatography. Affinity columns prepared with the GalNAc-specific Helix pomatia agglutinin (HPA) and equilibrated in THF specifically bind the (/sup 3/H)oligosaccharide derived from Forssman GL indicating that the immobilized HPA retained its carbohydrate-binding specificity in this solvent. Intact Forssman GL was bound by the HPA-column equilibrated in THF and was specifically eluted with 0.1 mg/ml GalNAc in THF. Purification of the Forssman GL was achieved when a crude lipid extract of sheep erythrocyte membranes was applied to the HPA-column in THF. Non-specifically bound GLs were eluted from the column using a step gradient of aqueous buffer in THF, while the addition of GalNAc was required to elute the specifically bound GLs. Using this procedure the A-active GLs were purified from a crude lipid extract of type A human erythrocytes in a single chromatographic step. The use of solvents that maintain carbohydrate-binding specificity and lipid solubility will permit the application of affinity chromatography on immobilized carbohydrate-binding proteins to intact GLs.

  2. Chemical Genomics Identifies the PERK-Mediated Unfolded Protein Stress Response as a Cellular Target for Influenza Virus Inhibition

    Directory of Open Access Journals (Sweden)

    Sara Landeras-Bueno

    2016-04-01

    Full Text Available Influenza A viruses generate annual epidemics and occasional pandemics of respiratory disease with important consequences for human health and the economy. Therefore, a large effort has been devoted to the development of new anti-influenza virus drugs directed to viral targets, as well as to the identification of cellular targets amenable to anti-influenza virus therapy. Here we have addressed the identification of such potential cellular targets by screening collections of drugs approved for human use. We reasoned that screening with a green fluorescent protein-based recombinant replicon system would identify cellular targets involved in virus transcription/replication and/or gene expression and hence address an early stage of virus infection. By using such a strategy, we identified Montelukast (MK as an inhibitor of virus multiplication. MK inhibited virus gene expression but did not alter viral RNA synthesis in vitro or viral RNA accumulation in vivo. The low selectivity index of MK prevented its use as an antiviral, but it was sufficient to identify a new cellular pathway suitable for anti-influenza virus intervention. By deep sequencing of RNA isolated from mock- and virus-infected human cells, treated with MK or left untreated, we showed that it stimulates the PERK-mediated unfolded protein stress response. The phosphorylation of PERK was partly inhibited in virus-infected cells but stimulated in MK-treated cells. Accordingly, pharmacological inhibition of PERK phosphorylation led to increased viral gene expression, while inhibition of PERK phosphatase reduced viral protein synthesis. These results suggest the PERK-mediated unfolded protein response as a potential cellular target to modulate influenza virus infection.

  3. In vitro evolution and affinity-maturation with Coliphage qβ display.

    Directory of Open Access Journals (Sweden)

    Claudia Skamel

    Full Text Available The Escherichia coli bacteriophage, Qβ (Coliphage Qβ, offers a favorable alternative to M13 for in vitro evolution of displayed peptides and proteins due to high mutagenesis rates in Qβ RNA replication that better simulate the affinity maturation processes of the immune response. We describe a benchtop in vitro evolution system using Qβ display of the VP1 G-H loop peptide of foot-and-mouth disease virus (FMDV. DNA encoding the G-H loop was fused to the A1 minor coat protein of Qβ resulting in a replication-competent hybrid phage that efficiently displayed the FMDV peptide. The surface-localized FMDV VP1 G-H loop cross-reacted with the anti-FMDV monoclonal antibody (mAb SD6 and was found to decorate the corners of the Qβ icosahedral shell by electron microscopy. Evolution of Qβ-displayed peptides, starting from fully degenerate coding sequences corresponding to the immunodominant region of VP1, allowed rapid in vitro affinity maturation to SD6 mAb. Qβ selected under evolutionary pressure revealed a non-canonical, but essential epitope for mAb SD6 recognition consisting of an Arg-Gly tandem pair. Finally, the selected hybrid phages induced polyclonal antibodies in guinea pigs with good affinity to both FMDV and hybrid Qβ-G-H loop, validating the requirement of the tandem pair epitope. Qβ-display emerges as a novel framework for rapid in vitro evolution with affinity-maturation to molecular targets.

  4. Purification of proteins specifically binding human endogenous retrovirus K long terminal repeat by affinity elution chromatography.

    Science.gov (United States)

    Trubetskoy, D O; Zavalova, L L; Akopov, S B; Nikolaev, L G

    2002-11-01

    A novel affinity elution procedure for purification of DNA-binding proteins was developed and employed to purify to near homogeneity the proteins recognizing a 21 base pair sequence within the long terminal repeat of human endogenous retroviruses K. The approach involves loading the initial protein mixture on a heparin-agarose column and elution of protein(s) of interest with a solution of double-stranded oligonucleotide containing binding sites of the protein(s). The affinity elution has several advantages over conventional DNA-affinity chromatography: (i) it is easier and faster, permitting to isolate proteins in a 1 day-one stage procedure; (ii) yield of a target protein is severalfold higher than that in DNA-affinity chromatography; (iii) it is not necessary to prepare a special affinity support for each factor to be isolated. Theaffinity elution could be a useful alternative to conventional DNA-affinity chromatography.

  5. Alternative affinity tools: more attractive than antibodies?

    NARCIS (Netherlands)

    Ruigrok, V.J.B.; Levisson, M.; Eppink, M.H.M.; Smidt, H.; Oost, van der J.

    2011-01-01

    Antibodies are the most successful affinity tools used today, in both fundamental and applied research (diagnostics, purification and therapeutics). Nonetheless, antibodies do have their limitations, including high production costs and low stability. Alternative affinity tools based on nucleic acids

  6. Spectral affinity in protein networks

    Directory of Open Access Journals (Sweden)

    Teng Shang-Hua

    2009-11-01

    Full Text Available Abstract Background Protein-protein interaction (PPI networks enable us to better understand the functional organization of the proteome. We can learn a lot about a particular protein by querying its neighborhood in a PPI network to find proteins with similar function. A spectral approach that considers random walks between nodes of interest is particularly useful in evaluating closeness in PPI networks. Spectral measures of closeness are more robust to noise in the data and are more precise than simpler methods based on edge density and shortest path length. Results We develop a novel affinity measure for pairs of proteins in PPI networks, which uses personalized PageRank, a random walk based method used in context-sensitive search on the Web. Our measure of closeness, which we call PageRank Affinity, is proportional to the number of times the smaller-degree protein is visited in a random walk that restarts at the larger-degree protein. PageRank considers paths of all lengths in a network, therefore PageRank Affinity is a precise measure that is robust to noise in the data. PageRank Affinity is also provably related to cluster co-membership, making it a meaningful measure. In our experiments on protein networks we find that our measure is better at predicting co-complex membership and finding functionally related proteins than other commonly used measures of closeness. Moreover, our experiments indicate that PageRank Affinity is very resilient to noise in the network. In addition, based on our method we build a tool that quickly finds nodes closest to a queried protein in any protein network, and easily scales to much larger biological networks. Conclusion We define a meaningful way to assess the closeness of two proteins in a PPI network, and show that our closeness measure is more biologically significant than other commonly used methods. We also develop a tool, accessible at http://xialab.bu.edu/resources/pnns, that allows the user to

  7. A shear-stress responsive nano-container to target critically constricted arteries

    OpenAIRE

    Holme, Margaret Nancy

    2014-01-01

    Atherosclerosis and associated cardiovascular diseases are the world's biggest cause of mortality. During the acute case of heart attack, vasodilators are administered to open up the constricted artery and allow blood perfusion to the surrounding tissue. However, there are currently no treatments on the market that allow such drugs to be delivered locally to the site of a critically constricted artery. Such a targeted delivery method could significantly improve patient prognosis. The presente...

  8. Identification of hookworm DAF-16/FOXO response elements and direct gene targets.

    Directory of Open Access Journals (Sweden)

    Xin Gao

    Full Text Available BACKGROUND: The infective stage of the parasitic nematode hookworm is developmentally arrested in the environment and needs to infect a specific host to complete its life cycle. The canine hookworm (Ancylostoma caninum is an excellent model for investigating human hookworm infections. The transcription factor of A. caninum, Ac-DAF-16, which has a characteristic fork head or "winged helix" DNA binding domain (DBD, has been implicated in the resumption of hookworm development in the host. However, the precise roles of Ac-DAF-16 in hookworm parasitism and its downstream targets are unknown. In the present study, we combined molecular techniques and bioinformatics to identify a group of Ac-DAF-16 binding sites and target genes. METHODOLOGY/PRINCIPAL FINDINGS: The DNA binding domain of Ac-DAF-16 was used to select genomic fragments by in vitro genomic selection. Twenty four bound genomic fragments were analyzed for the presence of the DAF-16 family binding element (DBE and possible alternative Ac-DAF-16 bind motifs. The 22 genes linked to these genomic fragments were identified using bioinformatics tools and defined as candidate direct gene targets of Ac-DAF-16. Their developmental stage-specific expression patterns were examined. Also, a new putative DAF-16 binding element was identified. CONCLUSIONS/SIGNIFICANCE: Our results show that Ac-DAF-16 is involved in diverse biological processes throughout hookworm development. Further investigation of these target genes will provide insights into the molecular basis by which Ac-DAF-16 regulates its downstream gene network in hookworm infection.

  9. Matrix Metalloprotease 2-Responsive Multifunctional Liposomal Nanocarrier for Enhanced Tumor Targeting

    Science.gov (United States)

    Zhu, Lin; Kate, Pooja; Torchilin, Vladimir P.

    2012-01-01

    A novel “smart” multifunctional drug delivery system was successfully developed to respond to the up-regulated matrix metalloprotease 2 (MMP2) in the tumor microenvironment and improve cancer cell-specific delivery of loaded drugs. The system represents a surface-functionalized liposomal nanocarrier, for which two functional polyethylene glycol (PEG)-lipid conjugates were synthesized and characterized. The functionalized liposome was further modified with the tumor cell-specific anti-nucleosome monoclonal antibody (mAb 2C5). In the resulting system, several drug delivery strategies were combined in the same nanocarrier in a simple way and coordinated in an optimal fashion. The functions of the nanocarrier include: i) the hydrophilic and flexible long PEG chains to prevent nanocarrier non-specific interactions and prolong its circulation time; ii) a nanoscale size of the system that allows for its passive tumor targeting via the enhanced permeability and retention (EPR) effect; iii) a mAb 2C5 to allow for the specific targeting of tumor cells; iv) a matrix metalloprotease 2-sensitive bond between PEG and lipid that undergoes cleavage in the tumor by the highly expressed extracellular MMP2 for the removal of PEG chains; v) The cell-penetrating peptide (TATp) triggering of the enhanced intracellular delivery of the system after long-chain PEG removal and exposure of the previously hidden surface-attached TATp. It is shown that such a design can enhance the targetability and internalization of nanocarriers in cancer cells. PMID:22409425

  10. pH-Responsive Core-Shell Structured Nanoparticles for Triple-Stage Targeted Delivery of Doxorubicin to Tumors.

    Science.gov (United States)

    Han, Lu; Tang, Cui; Yin, Chunhua

    2016-09-14

    The application of cytotoxic chemotherapeutics in cancer therapy has been largely restricted by their lack of selectivity. Despite the existence of numerous targeted delivery systems, it is practically challenging to develop one single system to simultaneously cover tumor-targeted delivery of chemotherapeutics at the tissue, cellular, and subcellular levels. To solve this problem, pH-responsive core-shell structured nanoparticles (CSNPs) were self-assembled in this study to provide triple-stage targeted delivery of doxorubicin (DOX) from the injection site to the nuclei of cancer cells. Amino-functionalized mesoporous silica nanoparticles (MSN) were doubly modified with TAT peptide and acid-cleavable polyethylene glycol (PEG) as the DOX-loaded cationic core. The anionic shell was constituted by galactose-modified poly(allylamine hydrochloride)-citraconic anhydride, a hepato-carcinoma-targeting polymer with charge-reversible property. In vitro results showed that PEG effectively reduced protein adsorption and phagocytic capture of CSNPs in the circulating blood (pH 7.4), thus facilitating passive accumulation in tumors (tissue level). Following PEG detachment via acidic hydrolysis in tumor microenvironment (pH 6.5), the exposed galactose ligands endowed CSNPs with active internalization into hepato-carcinoma cells (cellular level). Afterward, the acidity in endosomes and lysosomes (pH 5.0) triggered the conversion of anionic shell into positive charges, leading to core-shell disassembly and subsequent TAT-mediated delivery of DOX to the nuclei (subcellular level). Importantly, the efficiencies of each targeting moiety were nicely preserved when combining together in CSNPs. As a result, improved tumorous distribution and potent therapeutic efficacy of CSNPs were noted in tumor-bearing mice at a relatively low dose. CSNPs therefore provide an efficient and nontoxic platform for the targeted delivery of antitumor drugs. PMID:27558413

  11. Large-scale integration of small molecule-induced genome-wide transcriptional responses, Kinome-wide binding affinities and cell-growth inhibition profiles reveal global trends characterizing systems-level drug action

    Directory of Open Access Journals (Sweden)

    Dusica eVidovic

    2014-09-01

    Full Text Available The Library of Integrated Network-based Cellular Signatures (LINCS project is a large-scale coordinated effort to build a comprehensive systems biology reference resource. The goals of the program include the generation of a very large multidimensional data matrix and informatics and computational tools to integrate, analyze, and make the data readily accessible. LINCS data include genome-wide transcriptional signatures, biochemical protein binding profiles, cellular phenotypic response profiles and various other datasets for a wide range of cell model systems and molecular and genetic perturbations. Here we present a partial survey of this data facilitated by data standards and in particular a robust compound standardization workflow; we integrated several types of LINCS signatures and analyzed the results with a focus on mechanism of action and chemical compounds. We illustrate how kinase targets can be related to disease models and relevant drugs. We identified some fundamental trends that appear to link Kinome binding profiles and transcriptional signatures to chemical information and biochemical binding profiles to transcriptional responses independent of chemical similarity. To fill gaps in the datasets we developed and applied predictive models. The results can be interpreted at the systems level as demonstrated based on a large number of signaling pathways. We can identify clear global relationships, suggesting robustness of cellular responses to chemical perturbation. Overall, the results suggest that chemical similarity is a useful measure at the systems level, which would support phenotypic drug optimization efforts. With this study we demonstrate the potential of such integrated analysis approaches and suggest prioritizing further experiments to fill the gaps in the current data.

  12. Manifolds with integrable affine shape operator

    Directory of Open Access Journals (Sweden)

    Daniel A. Joaquín

    2005-05-01

    Full Text Available This work establishes the conditions for the existence of vector fields with the property that theirs covariant derivative, with respect to the affine normal connection, be the affine shape operatorS in hypersurfaces. Some results are obtained from this property and, in particular, for some kind of affine decomposable hypersurfaces we explicitely get the actual vector fields.

  13. Definition of the viral targets of protective HIV-1-specific T cell responses

    Directory of Open Access Journals (Sweden)

    Mothe Beatriz

    2011-12-01

    Full Text Available Abstract Background The efficacy of the CTL component of a future HIV-1 vaccine will depend on the induction of responses with the most potent antiviral activity and broad HLA class I restriction. However, current HIV vaccine designs are largely based on viral sequence alignments only, not incorporating experimental data on T cell function and specificity. Methods Here, 950 untreated HIV-1 clade B or -C infected individuals were tested for responses to sets of 410 overlapping peptides (OLP spanning the entire HIV-1 proteome. For each OLP, a "protective ratio" (PR was calculated as the ratio of median viral loads (VL between OLP non-responders and responders. Results For both clades, there was a negative relationship between the PR and the entropy of the OLP sequence. There was also a significant additive effect of multiple responses to beneficial OLP. Responses to beneficial OLP were of significantly higher functional avidity than responses to non-beneficial OLP. They also had superior in-vitro antiviral activities and, importantly, were at least as predictive of individuals' viral loads than their HLA class I genotypes. Conclusions The data thus identify immunogen sequence candidates for HIV and provide an approach for T cell immunogen design applicable to other viral infections.

  14. The ER mitochondria calcium cycle and ER stress response as therapeutic targets in amyotrophic lateral sclerosis

    Directory of Open Access Journals (Sweden)

    Vedrana eTadic

    2014-05-01

    Full Text Available Amyotrophic lateral sclerosis (ALS is a neurodegenerative disease characterized by progressive loss of upper and lower motor neurons. Although the etiology remains unclear, disturbances in calcium homoeostasis and protein folding are essential features of neurodegeneration in this disorder. Here, we review recent research findings on the interaction between endoplasmic reticulum (ER and mitochondria, and its effect on calcium signaling and oxidative stress. We further provide insights into studies, providing evidence that structures of the ER mitochondria calcium cycle (ERMCC serve as a promising targets for therapeutic approaches for treatment of ALS.

  15. Positive allosteric modulation of the GHB high-affinity binding site by the GABAA receptor modulator monastrol and the flavonoid catechin.

    Science.gov (United States)

    Eghorn, Laura F; Hoestgaard-Jensen, Kirsten; Kongstad, Kenneth T; Bay, Tina; Higgins, David; Frølund, Bente; Wellendorph, Petrine

    2014-10-01

    γ-Hydroxybutyric acid (GHB) is a metabolite of γ-aminobutyric acid (GABA) and a proposed neurotransmitter in the mammalian brain. We recently identified α4βδ GABAA receptors as possible high-affinity GHB targets. GABAA receptors are highly sensitive to allosteric modulation. Thus to investigate whether GHB high-affinity binding sites are also sensitive to allosteric modulation, we screened both known GABAA receptor ligands and a library of natural compounds in the rat cortical membrane GHB specific high-affinity [3H]NCS-382 binding assay. Two hits were identified: Monastrol, a positive allosteric modulator of GABA function at δ-containing GABAA receptors, and the naturally occurring flavonoid catechin. These compounds increased [3H]NCS-382 binding to 185-272% in high micromolar concentrations. Monastrol and (+)-catechin significantly reduced [3H]NCS-382 dissociation rates and induced conformational changes in the binding site, demonstrating a positive allosteric modulation of radioligand binding. Surprisingly, binding of [3H]GHB and the GHB high-affinity site-specific radioligands [125I]BnOPh-GHB and [3H]HOCPCA was either decreased or only weakly increased, indicating that the observed modulation was critically probe-dependent. Both monastrol and (+)-catechin were agonists at recombinant α4β3δ receptors expressed in Xenopus laevis oocytes. When monastrol and GHB were co-applied no changes were seen compared to the individual responses. In summary, we have identified the compounds monastrol and catechin as the first allosteric modulators of GHB high-affinity binding sites. Despite their relatively weak affinity, these compounds may aid in further characterization of the GHB high-affinity sites that are likely to represent certain GABAA receptors.

  16. A Borrelia burgdorferi Surface-Exposed Transmembrane Protein Lacking Detectable Immune Responses Supports Pathogen Persistence and Constitutes a Vaccine Target.

    Science.gov (United States)

    Kung, Faith; Kaur, Simarjot; Smith, Alexis A; Yang, Xiuli; Wilder, Cara N; Sharma, Kavita; Buyuktanir, Ozlem; Pal, Utpal

    2016-06-01

    Borrelia burgdorferi harbors a limited set of transmembrane surface proteins, most of which constitute key targets of humoral immune responses. Here we show that BB0405, a conserved membrane-spanning protein of unknown function, fails to evoke detectable antibody responses despite its extracellular exposure. bb0405 is a member of an operon and ubiquitously expressed throughout the rodent-tick infection cycle. The gene product serves an essential function in vivo, as bb0405-deletion mutants are unable to transmit from ticks and establish infection in mammalian hosts. Despite the lack of BB0405-specific immunoglobulin M or immunoglobulin G antibodies during natural infection, mice immunized with a recombinant version of the protein elicited high-titer and remarkably long-lasting antibody responses, conferring significant host protection against tick-borne infection. Taken together, these studies highlight the essential role of an apparently immune-invisible borrelial transmembrane protein in facilitating infection and its usefulness as a target of protective host immunity blocking the transmission of B. burgdorferi. PMID:26747708

  17. Hyaluronic acid-decorated graphene oxide nanohybrids as nanocarriers for targeted and pH-responsive anticancer drug delivery.

    Science.gov (United States)

    Song, Erqun; Han, Weiye; Li, Cheng; Cheng, Dan; Li, Lingrui; Liu, Lichao; Zhu, Guizhi; Song, Yang; Tan, Weihong

    2014-08-13

    A novel nanohybrid of hyaluronic acid (HA)-decorated graphene oxide (GO) was fabricated as a targeted and pH-responsive drug delivery system for controlling the release of anticancer drug doxorubicin (DOX) for tumor therapy. For the preparation, DOX was first loaded onto GO nanocarriers via π-π stacking and hydrogen-bonding interactions, and then it was decorated with HA to produce HA-GO-DOX nanohybrids via H-bonding interactions. In this strategy, HA served as both a targeting moiety and a hydrophilic group, making the as-prepared nanohybrids targeting, stable, and disperse. A high loading efficiency (42.9%) of DOX on the nanohybrids was also obtained. Cumulative DOX release from HA-GO-DOX was faster in pH 5.3 phosphate-buffered saline solution than that in pH 7.4, providing the basis for pH-response DOX release in the slightly acidic environment of tumor cells, while the much-slower DOX release from HA-GO-DOX than DOX showed the sustained drug-release capability of the nanohybrids. Fluorescent images of cellular uptake and cell viability analysis studies illustrated that these HA-GO-DOX nanohybrids significantly enhanced DOX accumulation in HA-targeted HepG2 cancer cells compared to HA-nontargeted RBMEC cells and subsequently induced selective cytotoxicity to HepG2 cells. In vivo antitumor efficiency of HA-GO-DOX nanohybrids showed obviously enhanced tumor inhibition rate for H22 hepatic cancer cell-bearing mice compared with free DOX and the GO-DOX formulation. These studies suggest that the HA-GO-DOX nanohybrids have potential clinical applications for anticancer drug delivery.

  18. Multivalent TB vaccines targeting the esx gene family generate potent and broad cell-mediated immune responses superior to BCG.

    Science.gov (United States)

    Villarreal, Daniel O; Walters, Jewell; Laddy, Dominick J; Yan, Jian; Weiner, David B

    2014-01-01

    Development of a broad-spectrum synthetic vaccine against TB would represent an important advance to the limited vaccine armamentarium against TB. It is believed that the esx family of TB antigens may represent important vaccine candidates. However, only 4 esx antigens have been studied as potential vaccine antigens. The challenge remains to develop a vaccine that simultaneously targets all 23 members of the esx family to induce enhanced broad-spectrum cell-mediated immunity. We sought to investigate if broader cellular immune responses could be induced using a multivalent DNA vaccine representing the esx family protein members delivered via electroporation. In this study, 15 designed esx antigens were created to cross target all members of the esx family. They were distributed into groups of 3 self-processing antigens each, resulting in 5 trivalent highly optimized DNA plasmids. Vaccination with all 5 constructs elicited robust antigen-specific IFN-γ responses to all encoded esx antigens and induced multifunctional CD4 Th1 and CD8 T cell responses. Importantly, we show that when all constructs are combined into a cocktail, the RSQ-15 vaccine, elicited substantial broad Ag-specific T cell responses to all esx antigens as compared with vaccination with BCG. Moreover, these vaccine-induced responses were highly cross-reactive with BCG encoded esx family members and were highly immune effective in a BCG DNA prime-boost format. Furthermore, we demonstrate the vaccine potential and immunopotent profile of several novel esx antigens never previously studied. These data highlight the likely importance of these novel immunogens for study as preventative or therapeutic synthetic TB vaccines in combination or as stand alone antigens.

  19. Acid-responsive PEGylated doxorubicin prodrug nanoparticles for neuropilin-1 receptor-mediated targeted drug delivery.

    Science.gov (United States)

    Song, Huijuan; Zhang, Ju; Wang, Weiwei; Huang, Pingsheng; Zhang, Yumin; Liu, Jianfeng; Li, Chen; Kong, Deling

    2015-12-01

    Self-assembled prodrug nanoparticles have demonstrated great promise in cancer chemotherapy. In the present study, we developed a new kind of prodrug nanoparticles for targeted drug delivery. PEGylated doxorubicin conjugate with an acid-cleavable cis-aconityl spacer was prepared. Then it was functionalized with a tumor-penetrating peptide, Cys-Arg-Gly-Asp-Lys (CRGDK), providing the prodrug nanoparticles with the specific binding ability to neurophilin-1 receptor. In acid mediums, doxorubicin could be released from the prodrug nanoparticles with an accumulative release around 60% through the acid-triggered hydrolysis of cis-aconityl bond and nanoparticle disassembly. Whereas, drug release was slow under a neutral pH and the accumulative drug release was less than 16%. In the cell culture tests, our prodrug nanoparticles showed enhanced endocytosis and cytotoxicity in cancer cells including HepG2, MCF-7 and MDA-MB-231 cells, but lower cytotoxicity in human cardiomyocyte H2C9. In the animal experiments, the prodrug nanoparticles were intravenously injected into Balb/c nude mice bearing MDA-MB-231 tumors. Enhanced drug penetration and accumulation in tumors, accompanying with a rapid early tumor-binding behavior, was observed after intravenous injection of the peptide modified prodrug nanoparticles. These data suggests that the acid-sensitive and tumor-targeting PEGylated doxorubicin prodrug nanoparticle may be an efficient drug delivery system for cancer chemotherapy.

  20. Identification of novel human damage response proteins targeted through yeast orthology.

    Directory of Open Access Journals (Sweden)

    J Peter Svensson

    Full Text Available Studies in Saccharomyces cerevisiae show that many proteins influence cellular survival upon exposure to DNA damaging agents. We hypothesized that human orthologs of these S. cerevisiae proteins would also be required for cellular survival after treatment with DNA damaging agents. For this purpose, human homologs of S. cerevisiae proteins were identified and mapped onto the human protein-protein interaction network. The resulting human network was highly modular and a series of selection rules were implemented to identify 45 candidates for human toxicity-modulating proteins. The corresponding transcripts were targeted by RNA interference in human cells. The cell lines with depleted target expression were challenged with three DNA damaging agents: the alkylating agents MMS and 4-NQO, and the oxidizing agent t-BuOOH. A comparison of the survival revealed that the majority (74% of proteins conferred either sensitivity or resistance. The identified human toxicity-modulating proteins represent a variety of biological functions: autophagy, chromatin modifications, RNA and protein metabolism, and telomere maintenance. Further studies revealed that MMS-induced autophagy increase the survival of cells treated with DNA damaging agents. In summary, we show that damage recovery proteins in humans can be identified through homology to S. cerevisiae and that many of the same pathways are represented among the toxicity modulators.

  1. Elucidating induced plant defenses: the use of targeted metabolomics as a bridge from elicitation to response

    Science.gov (United States)

    Dynamic plant defense responses to biotic attack involve the perception of specific biochemical elicitors associated with the offending agent, activation of signaling cascades, and the production of small molecules with complex protective roles. Chemical analyses are essential empirical tools for el...

  2. Early Detection Rapid Response Program Targets New Noxious Weed Species in Washington State

    Science.gov (United States)

    Andreas, Jennifer E.; Halpern, Alison D.; DesCamp, Wendy C.; Miller, Timothy W.

    2015-01-01

    Early detection, rapid response is a critical component of invasive plant management. It can be challenging, however, to detect new invaders before they become established if landowners cannot identify species of concern. In order to increase awareness, eye-catching postcards were developed in Washington State as part of a noxious weed educational…

  3. Specific nongluten proteins of wheat are novel target antigens in celiac disease humoral response

    Science.gov (United States)

    Background: Celiac disease is an immune-mediated enteropathy that is generally understood to be triggered by the ingestion of gluten proteins of wheat and related cereals. The skin manifestation of the condition is known as dermatitis herpetiformis. Antibody response to native and deamidated seque...

  4. Transcriptional profiling uncovers a network of cholesterol-responsive atherosclerosis target genes.

    Directory of Open Access Journals (Sweden)

    Josefin Skogsberg

    2008-03-01

    Full Text Available Despite the well-documented effects of plasma lipid lowering regimes halting atherosclerosis lesion development and reducing morbidity and mortality of coronary artery disease and stroke, the transcriptional response in the atherosclerotic lesion mediating these beneficial effects has not yet been carefully investigated. We performed transcriptional profiling at 10-week intervals in atherosclerosis-prone mice with human-like hypercholesterolemia and a genetic switch to lower plasma lipoproteins (Ldlr(-/-Apo(100/100Mttp(flox/flox Mx1-Cre. Atherosclerotic lesions progressed slowly at first, then expanded rapidly, and plateaued after advanced lesions formed. Analysis of lesion expression profiles indicated that accumulation of lipid-poor macrophages reached a point that led to the rapid expansion phase with accelerated foam-cell formation and inflammation, an interpretation supported by lesion histology. Genetic lowering of plasma cholesterol (e.g., lipoproteins at this point all together prevented the formation of advanced plaques and parallel transcriptional profiling of the atherosclerotic arterial wall identified 37 cholesterol-responsive genes mediating this effect. Validation by siRNA-inhibition in macrophages incubated with acetylated-LDL revealed a network of eight cholesterol-responsive atherosclerosis genes regulating cholesterol-ester accumulation. Taken together, we have identified a network of atherosclerosis genes that in response to plasma cholesterol-lowering prevents the formation of advanced plaques. This network should be of interest for the development of novel atherosclerosis therapies.

  5. Complement receptor type two (CR2,CR21): a target for influencing the humoral immune response and antigen-trapping.

    Science.gov (United States)

    Prodinger, W M

    1999-01-01

    Cellular receptors for complement C3 fragments deposited on antigens are important bricks in the wall defending against microbial pathogens. The part of complement receptor type 2 (CR2; CD21) deals with enhancing humoral immune responses and with long-term trapping of C3d-coated antigen by follicular dendritic cells. CR2 is also pivotal for Epstein-Barr virus (EBV) infection. Here, the current understanding, how CR2 interacts with its ligands C3d, EBV, and CD23 is summarized. The potential to target CR2 for clinical therapy or immunization purposes are discussed. PMID:10741859

  6. Clinical Response of Metastatic Breast Cancer to Multi-targeted Therapeutic Approach: A Single Case Report

    Directory of Open Access Journals (Sweden)

    Christian Meiners

    2011-03-01

    Full Text Available The present article describes the ongoing (partial remission of a female patient (41 years old from estrogen receptor (ER-positive/progesterone receptor (PR-negative metastatic breast cancer in response to a combination treatment directed towards the revitalization of the mitochondrial respiratory chain (oxidative phosphorylation, the suppression of NF-kappaB as a factor triggering the inflammatory response, and chemotherapy with capecitabine. The reduction of tumor mass was evidenced by a continuing decline of CA15-3 and CEA tumor marker serum levels and 18FDG-PET-CT plus magnetic resonance (MR imaging. It is concluded that such combination treatment might be a useful option for treating already formed metastases and for providing protection against the formation of metastases in ER positive breast cancer. The findings need to be corroborated by clinical trials. Whether similar results can be expected for other malignant tumor phenotypes relying on glycolysis as the main energy source remains to be elucidated.

  7. Clinical Response of Metastatic Breast Cancer to Multi-targeted Therapeutic Approach: A Single Case Report

    Energy Technology Data Exchange (ETDEWEB)

    Meiners, Christian [Gautinger Straße 3b, D-82234 Wessling (Germany)

    2011-03-17

    The present article describes the ongoing (partial) remission of a female patient (41 years old) from estrogen receptor (ER)-positive/progesterone receptor (PR)-negative metastatic breast cancer in response to a combination treatment directed towards the revitalization of the mitochondrial respiratory chain (oxidative phosphorylation), the suppression of NF-kappaB as a factor triggering the inflammatory response, and chemotherapy with capecitabine. The reduction of tumor mass was evidenced by a continuing decline of CA15-3 and CEA tumor marker serum levels and {sup 18}FDG-PET-CT plus magnetic resonance (MR) imaging. It is concluded that such combination treatment might be a useful option for treating already formed metastases and for providing protection against the formation of metastases in ER positive breast cancer. The findings need to be corroborated by clinical trials. Whether similar results can be expected for other malignant tumor phenotypes relying on glycolysis as the main energy source remains to be elucidated.

  8. Regulation of MicroRNA-155 in Atherosclerotic Inflammatory Responses by Targeting MAP3K10

    OpenAIRE

    Zhu, Jianhua; Chen, Ting; Yang, Lin; Li, Zhoubin; Wong, Mei Mei; Zheng, Xiaoye; Pan, Xiaoping; Zhang, Li; Yan, Hui

    2012-01-01

    Aims Accumulating evidence suggest that numerous microRNAs (miRNAs) play important roles in cell proliferation, apoptosis, and differentiation, as well as various diseases that accompany inflammatory responses. Inflammation is known to be a major contributor to atherogenesis. Previous studies provide promising evidence in support of the role of miRNAs in cardiovascular disease. However, mechanistic data on these small molecules in atherosclerosis (AS) are still missing. The present study aims...

  9. Control of target gene specificity during metamorphosis by the steroid response gene E93

    OpenAIRE

    Mou, Xiaochun; Duncan, Dianne M.; Baehrecke, Eric H.; Duncan, Ian

    2012-01-01

    Hormonal control of sexual maturation is a common feature in animal development. A particularly dramatic example is the metamorphosis of insects, in which pulses of the steroid hormone ecdysone drive the wholesale transformation of the larva into an adult. The mechanisms responsible for this transformation are not well understood. Work in Drosophila indicates that the larval and adult forms are patterned by the same underlying sets of developmental regulators, but it is not understood how the...

  10. Ligand-responsive RNA switches: viral translation regulators, therapeutic targets, and tunable building blocks for nanotechnology

    OpenAIRE

    Boerneke, Mark A.

    2016-01-01

    Ligand-responsive RNA mechanical switches represent a new class of simple and small switching modules which regulate viral translation initiation by adopting well-defined ligand-free and bound conformational states without undergoing large secondary structure rearrangements, distinguishing them from metabolite-sensing riboswitches. Initially discovered in the internal ribosome entry site (IRES) of hepatitis C virus (HCV), RNA switch motifs have now been discovered in the genomes of diverse o...

  11. Rational self-affine tiles

    CERN Document Server

    Steiner, Wolfgang

    2012-01-01

    An integral self-affine tile is the solution of a set equation $\\mathbf{A} \\mathcal{T} = \\bigcup_{d \\in \\mathcal{D}} (\\mathcal{T} + d)$, where $\\mathbf{A}$ is an $n \\times n$ integer matrix and $\\mathcal{D}$ is a finite subset of $\\mathbb{Z}^n$. In the recent decades, these objects and the induced tilings have been studied systematically. We extend this theory to matrices $\\mathbf{A} \\in \\mathbb{Q}^{n \\times n}$. We define rational self-affine tiles as compact subsets of the open subring $\\mathbb{R}^n\\times \\prod_\\mathfrak{p} K_\\mathfrak{p}$ of the ad\\'ele ring $\\mathbb{A}_K$, where the factors of the (finite) product are certain $\\mathfrak{p}$-adic completions of a number field $K$ that is defined in terms of the characteristic polynomial of $\\mathbf{A}$. Employing methods from classical algebraic number theory, Fourier analysis in number fields, and results on zero sets of transfer operators, we establish a general tiling theorem for these tiles. We also associate a second kind of tiles with a rational matr...

  12. Graded electron affinity electron source

    Energy Technology Data Exchange (ETDEWEB)

    Shaw, J.L.; Gray, H.F.; Jensen, K.L.; Jung, T.M. [Naval Research Laboratory, Washington, DC 20375 (United States)

    1996-05-01

    We describe a new electron source using electric field and low electron affinity semiconductor materials to bring charge to potential energy levels near the vacuum level while still in the solid. The basic idea involves moving some of the potential barrier from the surface to the bulk, and distributing the barrier over a thin layer below the surface. In so doing, the emission physics is changed fundamentally from a quantum mechanical tunneling process largely controlled by surface properties to a classical transport process largely controlled by the band structure of a wide bandgap semiconductor. The composition of the thin layer below the surface would be graded such that the conduction band minimum changes from an energy close to the substrate Fermi level to an energy significantly closer to the vacuum level. Electrons from the substrate would be drawn into the graded composition layer with an electric field produced by a pointed emitter structure and extraction gate similar to that used in field emitter arrays. Relative to a conventional field emitter array, the new source is expected to require lower extraction fields and exhibit improved emission uniformity, stability, and noise. The {ital I}{endash}{ital V} characteristics are not Fowler{endash}Nordheim and may include a saturation effect at high current densities that could be engineered to improve reliability and uniformity. The group III nitrides can be grown with a continuous range of composition that provides the range of electron affinity needed to produce the proposed electronic structure. {copyright} {ital 1996 American Vacuum Society}

  13. The affine quantum gravity programme

    International Nuclear Information System (INIS)

    The central principle of affine quantum gravity is securing and maintaining the strict positivity of the matrix { g-hat ab(x)} composed of the spatial components of the local metric operator. On spectral grounds, canonical commutation relations are incompatible with this principle, and they must be replaced by noncanonical, affine commutation relations. Due to the partial second-class nature of the quantum gravitational constraints, it is advantageous to use the recently developed projection operator method, which treats all quantum constraints on an equal footing. Using this method, enforcement of regularized versions of the gravitational operator constraints is formulated quite naturally by means of a novel and relatively well-defined functional integral involving only the same set of variables that appears in the usual classical formulation. It is anticipated that skills and insight to study this formulation can be developed by studying special, reduced-variable models that still retain some basic characteristics of gravity, specifically a partial second-class constraint operator structure. Although perturbatively nonrenormalizable, gravity may possibly be understood nonperturbatively from a hard-core perspective that has proved valuable for specialized models. Finally, developing a procedure to pass to the genuine physical Hilbert space involves several interconnected steps that require careful coordination

  14. Development of an aptamer-based affinity purification method for vascular endothelial growth factor

    Directory of Open Access Journals (Sweden)

    Maren Lönne

    2015-12-01

    Full Text Available Since aptamers bind their targets with high affinity and specificity, they are promising alternative ligands in protein affinity purification. As aptamers are chemically synthesized oligonucleotides, they can be easily produced in large quantities regarding GMP conditions allowing their application in protein production for therapeutic purposes. Several advantages of aptamers compared to antibodies are described in general within this paper. Here, an aptamer directed against the human Vascular Endothelial Growth Factor (VEGF was used as affinity ligand for establishing a purification platform for VEGF in small scale. The aptamer was covalently immobilized on magnetic beads in a controlled orientation resulting in a functional active affinity matrix. Target binding was optimized by introduction of spacer molecules and variation of aptamer density. Further, salt-induced target elution was demonstrated as well as VEGF purification from a complex protein mixture proving the specificity of protein-aptamer binding.

  15. The intestinal T cell response to alpha-gliadin in adult celiac disease is focused on a single deamidated glutamine targeted by tissue transglutaminase

    DEFF Research Database (Denmark)

    Arentz-Hansen, H; Körner, R; Molberg, O;

    2000-01-01

    The great majority of patients that are intolerant of wheat gluten protein due to celiac disease (CD) are human histocompatibility leukocyte antigen (HLA)-DQ2(+), and the remaining few normally express HLA-DQ8. These two class II molecules are chiefly responsible for the presentation of gluten...... for T cell recognition. Gluten-specific T cell lines from 16 different adult patients all responded to one or both of these deamidated peptides, indicating that these epitopes are highly relevant to disease pathology. Binding studies showed that the deamidated peptides displayed an increased affinity...

  16. Complementary three-dimensional quantitative structure-activity relationship modeling of binding affinity and functional potency

    DEFF Research Database (Denmark)

    Tosco, Paolo; Ahring, Philip K; Dyhring, Tino;

    2009-01-01

    Complementary 3D-QSAR modeling of binding affinity and functional potency is proposed as a tool to pinpoint the molecular features of the ligands, and the corresponding amino acids in the receptor, responsible for high affinity binding vs those driving agonist behavior and receptor activation...

  17. Defining a Radiomic Response Phenotype: A Pilot Study using targeted therapy in NSCLC.

    Science.gov (United States)

    Aerts, Hugo J W L; Grossmann, Patrick; Tan, Yongqiang; Oxnard, Geoffrey G; Rizvi, Naiyer; Schwartz, Lawrence H; Zhao, Binsheng

    2016-01-01

    Medical imaging plays a fundamental role in oncology and drug development, by providing a non-invasive method to visualize tumor phenotype. Radiomics can quantify this phenotype comprehensively by applying image-characterization algorithms, and may provide important information beyond tumor size or burden. In this study, we investigated if radiomics can identify a gefitinib response-phenotype, studying high-resolution computed-tomography (CT) imaging of forty-seven patients with early-stage non-small cell lung cancer before and after three weeks of therapy. On the baseline-scan, radiomic-feature Laws-Energy was significantly predictive for EGFR-mutation status (AUC = 0.67, p = 0.03), while volume (AUC = 0.59, p = 0.27) and diameter (AUC = 0.56, p = 0.46) were not. Although no features were predictive on the post-treatment scan (p > 0.08), the change in features between the two scans was strongly predictive (significant feature AUC-range = 0.74-0.91). A technical validation revealed that the associated features were also highly stable for test-retest (mean ± std: ICC = 0.96 ± 0.06). This pilot study shows that radiomic data before treatment is able to predict mutation status and associated gefitinib response non-invasively, demonstrating the potential of radiomics-based phenotyping to improve the stratification and response assessment between tyrosine kinase inhibitors (TKIs) sensitive and resistant patient populations. PMID:27645803

  18. Defining a Radiomic Response Phenotype: A Pilot Study using targeted therapy in NSCLC

    Science.gov (United States)

    Aerts, Hugo J. W. L.; Grossmann, Patrick; Tan, Yongqiang; Oxnard, Geoffrey G.; Rizvi, Naiyer; Schwartz, Lawrence H.; Zhao, Binsheng

    2016-01-01

    Medical imaging plays a fundamental role in oncology and drug development, by providing a non-invasive method to visualize tumor phenotype. Radiomics can quantify this phenotype comprehensively by applying image-characterization algorithms, and may provide important information beyond tumor size or burden. In this study, we investigated if radiomics can identify a gefitinib response-phenotype, studying high-resolution computed-tomography (CT) imaging of forty-seven patients with early-stage non-small cell lung cancer before and after three weeks of therapy. On the baseline-scan, radiomic-feature Laws-Energy was significantly predictive for EGFR-mutation status (AUC = 0.67, p = 0.03), while volume (AUC = 0.59, p = 0.27) and diameter (AUC = 0.56, p = 0.46) were not. Although no features were predictive on the post-treatment scan (p > 0.08), the change in features between the two scans was strongly predictive (significant feature AUC-range = 0.74–0.91). A technical validation revealed that the associated features were also highly stable for test-retest (mean ± std: ICC = 0.96 ± 0.06). This pilot study shows that radiomic data before treatment is able to predict mutation status and associated gefitinib response non-invasively, demonstrating the potential of radiomics-based phenotyping to improve the stratification and response assessment between tyrosine kinase inhibitors (TKIs) sensitive and resistant patient populations. PMID:27645803

  19. An affinity pull-down approach to identify the plant cyclic nucleotide interactome

    KAUST Repository

    Donaldson, Lara Elizabeth

    2013-09-03

    Cyclic nucleotides (CNs) are intracellular second messengers that play an important role in mediating physiological responses to environmental and developmental signals, in species ranging from bacteria to humans. In response to these signals, CNs are synthesized by nucleotidyl cyclases and then act by binding to and altering the activity of downstream target proteins known as cyclic nucleotide-binding proteins (CNBPs). A number of CNBPs have been identified across kingdoms including transcription factors, protein kinases, phosphodiesterases, and channels, all of which harbor conserved CN-binding domains. In plants however, few CNBPs have been identified as homology searches fail to return plant sequences with significant matches to known CNBPs. Recently, affinity pull-down techniques have been successfully used to identify CNBPs in animals and have provided new insights into CN signaling. The application of these techniques to plants has not yet been extensively explored and offers an alternative approach toward the unbiased discovery of novel CNBP candidates in plants. Here, an affinity pull-down technique for the identification of the plant CN interactome is presented. In summary, the method involves an extraction of plant proteins which is incubated with a CN-bait, followed by a series of increasingly stringent elutions that eliminates proteins in a sequential manner according to their affinity to the bait. The eluted and bait-bound proteins are separated by one-dimensional gel electrophoresis, excised, and digested with trypsin after which the resultant peptides are identified by mass spectrometry - techniques that are commonplace in proteomics experiments. The discovery of plant CNBPs promises to provide valuable insight into the mechanism of CN signal transduction in plants. © Springer Science+Business Media New York 2013.

  20. The Dietary Flavonoid Kaempferol Mediates Anti-Inflammatory Responses via the Src, Syk, IRAK1, and IRAK4 Molecular Targets

    Directory of Open Access Journals (Sweden)

    Shi Hyoung Kim

    2015-01-01

    Full Text Available Even though a lot of reports have suggested the anti-inflammatory activity of kaempferol (KF in macrophages, little is known about its exact anti-inflammatory mode of action and its immunopharmacological target molecules. In this study, we explored anti-inflammatory activity of KF in LPS-treated macrophages. In particular, molecular targets for KF action were identified by using biochemical and molecular biological analyses. KF suppressed the release of nitric oxide (NO and prostaglandin E2 (PGE2, downregulated the cellular adhesion of U937 cells to fibronectin (FN, neutralized the generation of radicals, and diminished mRNA expression levels of inflammatory genes encoding inducible NO synthase (iNOS, TNF-α, and cyclooxygenase- (COX- 2 in lipopolysaccharide- (LPS- and sodium nitroprusside- (SNP- treated RAW264.7 cells and peritoneal macrophages. KF reduced NF-κB (p65 and p50 and AP-1 (c-Jun and c-Fos levels in the nucleus and their transcriptional activity. Interestingly, it was found that Src, Syk, IRAK1, and IRAK4 responsible for NF-κB and AP-1 activation were identified as the direct molecular targets of KF by kinase enzyme assays and by measuring their phosphorylation patterns. KF was revealed to have in vitro and in vivo anti-inflammatory activity by the direct suppression of Src, Syk, IRAK1, and IRAK4, involved in the activation of NF-κB and AP-1.

  1. microRNAs targeting DEAD-box helicases are involved in salinity stress response in rice (Oryza sativa L.

    Directory of Open Access Journals (Sweden)

    Macovei Anca

    2012-10-01

    Full Text Available Abstract Background Rice (Oryza sativa L., one of the most important food crop in the world, is considered to be a salt-sensitive crop. Excess levels of salt adversely affect all the major metabolic activities, including cell wall damage, cytoplasmic lysis and genomic stability. In order to cope with salt stress, plants have evolved high degrees of developmental plasticity, including adaptation via cascades of molecular networks and changes in gene expression profiles. Posttranscriptional regulation, through the activity of microRNAs, also plays an important role in the plant response to salinity conditions. MicroRNAs are small endogenous RNAs that modulate gene expression and are involved in the most essential physiological processes, including plant development and adaptation to environmental changes. Results In the present study, we investigated the expression profiles of osa-MIR414, osa-MIR408 and osa-MIR164e along with their targeted genes, under salinity stress conditions in wild type and transgenic rice plants ectopically expressing the PDH45 (Pea DNA Helicase gene. The present miRNAs were predicted to target the OsABP (ATP-Binding Protein, OsDSHCT (DOB1/SK12/helY-like DEAD-box Helicase and OsDBH (DEAD-Box Helicase genes, included in the DEAD-box helicase family. An in silico characterization of the proteins was performed and the miRNAs predicted targets were validated by RLM-5′RACE. The qRT-PCR analysis showed that the OsABP, OsDBH and OsDSHCT genes were up-regulated in response to 100 and 200 mM NaCl treatments. The present study also highlighted an increased accumulation of the gene transcripts in wild type plants, with the exception of the OsABP mRNA which showed the highest level (15.1-fold change compared to control in the transgenic plants treated with 200 mM NaCl. Salinity treatments also affected the expression of osa-MIR414, osa-MIR164e and osa-MIR408, found to be significantly down-regulated, although the changes in mi

  2. Receptor-targeted metalloradiopharmaceuticals. Final technical report

    International Nuclear Information System (INIS)

    Copper (II) and platinum (II) coordination complexes were prepared and characterized. These complexes were designed to afford structural homology with steroidal and non-steroidal estrogens for possible use as receptor-targeted radiopharmaceuticals. While weak affinity for the estrogen receptor was detectable, none would appear to have sufficient receptor-affinity for estrogen-receptor-targeted imaging or therapy

  3. BCL-2 family proteins as 5-Azacytidine-sensitizing targets and determinants of response in myeloid malignancies

    Science.gov (United States)

    Bogenberger, J M; Kornblau, S M; Pierceall, W E; Lena, R; Chow, D; Shi, C-X; Mantei, J; Ahmann, G; Gonzales, I M; Choudhary, A; Valdez, R; Camoriano, J; Fauble, V; Tiedemann, R E; Qiu, Y H; Coombes, K R; Cardone, M; Braggio, E; Yin, H; Azorsa, D O; Mesa, R A; Stewart, A K; Tibes, R

    2014-01-01

    Synergistic molecular vulnerabilities enhancing hypomethylating agents in myeloid malignancies have remained elusive. RNA-interference drug modifier screens identified antiapoptotic BCL-2 family members as potent 5-Azacytidine-sensitizing targets. In further dissecting BCL-XL, BCL-2 and MCL-1 contribution to 5-Azacytidine activity, siRNA silencing of BCL-XL and MCL-1, but not BCL-2, exhibited variable synergy with 5-Azacytidine in vitro. The BCL-XL, BCL-2 and BCL-w inhibitor ABT-737 sensitized most cell lines more potently compared with the selective BCL-2 inhibitor ABT-199, which synergized with 5-Azacytidine mostly at higher doses. Ex vivo, ABT-737 enhanced 5-Azacytidine activity across primary AML, MDS and MPN specimens. Protein levels of BCL-XL, BCL-2 and MCL-1 in 577 AML patient samples showed overlapping expression across AML FAB subtypes and heterogeneous expression within subtypes, further supporting a concept of dual/multiple BCL-2 family member targeting consistent with RNAi and pharmacologic results. Consequently, silencing of MCL-1 and BCL-XL increased the activity of ABT-199. Functional interrogation of BCL-2 family proteins by BH3 profiling performed on patient samples significantly discriminated clinical response versus resistance to 5-Azacytidine-based therapies. On the basis of these results, we propose a clinical trial of navitoclax (clinical-grade ABT-737) combined with 5-Azacytidine in myeloid malignancies, as well as to prospectively validate BH3 profiling in predicting 5-Azacytidine response. PMID:24451410

  4. BCL-2 family proteins as 5-Azacytidine-sensitizing targets and determinants of response in myeloid malignancies.

    Science.gov (United States)

    Bogenberger, J M; Kornblau, S M; Pierceall, W E; Lena, R; Chow, D; Shi, C-X; Mantei, J; Ahmann, G; Gonzales, I M; Choudhary, A; Valdez, R; Camoriano, J; Fauble, V; Tiedemann, R E; Qiu, Y H; Coombes, K R; Cardone, M; Braggio, E; Yin, H; Azorsa, D O; Mesa, R A; Stewart, A K; Tibes, R

    2014-08-01

    Synergistic molecular vulnerabilities enhancing hypomethylating agents in myeloid malignancies have remained elusive. RNA-interference drug modifier screens identified antiapoptotic BCL-2 family members as potent 5-Azacytidine-sensitizing targets. In further dissecting BCL-XL, BCL-2 and MCL-1 contribution to 5-Azacytidine activity, siRNA silencing of BCL-XL and MCL-1, but not BCL-2, exhibited variable synergy with 5-Azacytidine in vitro. The BCL-XL, BCL-2 and BCL-w inhibitor ABT-737 sensitized most cell lines more potently compared with the selective BCL-2 inhibitor ABT-199, which synergized with 5-Azacytidine mostly at higher doses. Ex vivo, ABT-737 enhanced 5-Azacytidine activity across primary AML, MDS and MPN specimens. Protein levels of BCL-XL, BCL-2 and MCL-1 in 577 AML patient samples showed overlapping expression across AML FAB subtypes and heterogeneous expression within subtypes, further supporting a concept of dual/multiple BCL-2 family member targeting consistent with RNAi and pharmacologic results. Consequently, silencing of MCL-1 and BCL-XL increased the activity of ABT-199. Functional interrogation of BCL-2 family proteins by BH3 profiling performed on patient samples significantly discriminated clinical response versus resistance to 5-Azacytidine-based therapies. On the basis of these results, we propose a clinical trial of navitoclax (clinical-grade ABT-737) combined with 5-Azacytidine in myeloid malignancies, as well as to prospectively validate BH3 profiling in predicting 5-Azacytidine response. PMID:24451410

  5. Molecular targets in cellular response to ionizing radiation and implications in space radiation protection

    Energy Technology Data Exchange (ETDEWEB)

    Belli, M.; Tabocchini, M.A. [Istituto Superiore di Sanita, Rome (Italy). Physics Lab.; Sapora, O. [Istituto Superiore di Sanita, Rome (Italy). Comparative Toxicology Lab.

    2002-12-01

    DNA repair systems and cell cycle checkpoints closely co-operate in the attempt of maintaining the genomic integrity of cells damaged by ionizing radiation. DNA double-strand breaks (DSB) are considered as the most biologically important radiation-induced damage. Their spatial distribution and association with other types of damage depend on radiation quality. It is believed these features affect damage reparability, thus explaining the higher efficiency for cellular effects of densely ionizing radiation with respect to {gamma}-rays. DSB repair systems identified in mammalian cells are homologous recombination (HR), single-strand annealing (SSA) and non-homologous end-joining (NHEJ). Some enzymes may participate in more than one of these repair systems. DNA damage also triggers biochemical signals activating checkpoints responsible for delay in cell cycle progression that allows more time for repair. Those at G1/S and S phases prevent replication of damaged DNA and those at G2/M phase prevent segregation of changed chromosomes. Individuals with lack or alterations of genes involved in DNA DSB repair and cell cycle checkpoints exhibit syndromes characterized by genome instability and predisposition to cancer. Information reviewed in this paper on the basic mechanisms of cellular response to ionizing radiation indicates their importance for a number of issues relevant to protection of astronauts from space radiation. (author)

  6. Novel trends in affinity biosensors: current challenges and perspectives

    International Nuclear Information System (INIS)

    Molecular biorecognition processes facilitate physical and biochemical interactions between molecules in all crucial metabolic pathways. Perhaps the target analyte and the biorecognition element interactions have the most impactful use in biosensing applications. Traditional analytical sensing systems offer excellent biorecognition elements with the ability to detect and determine the presence of analytes. High affinity antibodies and DNA play an important role in the development of affinity biosensors based on electrochemical, optical and mass sensitive approaches. Advancements in this area routinely employ labels, label free, nanoparticles, multifunctional matrices, carbon nanotubes and other methods to meet the requirements of its own application. However, despite increasing affinity ceilings for conventional biosensors, the field draws back in meeting specifically important demands, such as long-term stability, ultrasensitivity, rapid detection, extreme selectivity, strong biological base, calibration, in vivo measurements, regeneration, satisfactory performance and ease of production. Nevertheless, recent efforts through this line have produced novel high-tech nanosensing systems such as ‘aptamers’ and ‘phages’ which exhibit high-throughput sensing. Aptamers and phages are powerful tools that excel over antibodies in sensibility, stability, multi-detection, in vivo measurements and regeneration. Phages are superior in stability, screening for affinity-based target molecules ranging from small to proteins and even cells, and easy production. In this review, we focus mainly on recent developments in affinity-based biosensors such as immunosensors, DNA sensors, emphasizing aptasensors and phage-based biosensors basing on novel electrochemical, optical and mass sensitive detection techniques. We also address enzyme inhibition-based biosensors and the current problems associated with the above sensors and their future perspectives. (topical review)

  7. Targeting of Antibodies using Aptamers

    OpenAIRE

    Missailidis, Sotiris

    2003-01-01

    The chapter presents a methodology for the rapid selection of aptamers against antibody targets. It is a detailed account of the various methodological steps that describe the selection of aptamers, including PCR steps, buffers to be used, target immobilisation, partitioning and amplification of aptamers, clonning and sequencing, to results in high affinity and specificity ligands for the chosen target antibody.

  8. Pulmonary fibrosis in response to environmental cues and molecular targets involved in its pathogenesis.

    Science.gov (United States)

    Yoshida, Toshinori; Ohnuma, Aya; Horiuchi, Haruka; Harada, Takanori

    2011-03-01

    Chronic lung injury resulting from a variety of different causes is frequently associated with the develop ment of pulmonary fibrosis in humans. Although the etiology of pulmonary fibrosis is generally unknown, several sources of evidence support the hypothesis that a number of environmental and occupational agents play an etiologic role in the pathogenesis of this disease. The agents discussed in this review include beryllium, nylon flock, textile printing aerosols, polyvinyl chloride and didecyldimethylammonium chloride. The authors also describe a variety of animal models, including genetically modified mice, in order to investigate the molecular mechanism of pulmonary fibrosis, focusing on chemokine receptors, regulatory T cells and transforming growth factor-β and bone morphogenetic protein signaling. Overall, we propose the concept of toxicological pulmonary fibrosis as a lung disease induced in response to environmental cues.

  9. Aspirin's Active Metabolite Salicylic Acid Targets High Mobility Group Box 1 to Modulate Inflammatory Responses.

    Science.gov (United States)

    Choi, Hyong Woo; Tian, Miaoying; Song, Fei; Venereau, Emilie; Preti, Alessandro; Park, Sang-Wook; Hamilton, Keith; Swapna, G V T; Manohar, Murli; Moreau, Magali; Agresti, Alessandra; Gorzanelli, Andrea; De Marchis, Francesco; Wang, Huang; Antonyak, Marc; Micikas, Robert J; Gentile, Daniel R; Cerione, Richard A; Schroeder, Frank C; Montelione, Gaetano T; Bianchi, Marco E; Klessig, Daniel F

    2015-01-01

    Salicylic acid (SA) and its derivatives have been used for millennia to reduce pain, fever and inflammation. In addition, prophylactic use of acetylsalicylic acid, commonly known as aspirin, reduces the risk of heart attack, stroke and certain cancers. Because aspirin is rapidly de-acetylated by esterases in human plasma, much of aspirin's bioactivity can be attributed to its primary metabolite, SA. Here we demonstrate that human high mobility group box 1 (HMGB1) is a novel SA-binding protein. SA-binding sites on HMGB1 were identified in the HMG-box domains by nuclear magnetic resonance (NMR) spectroscopic studies and confirmed by mutational analysis. Extracellular HMGB1 is a damage-associated molecular pattern molecule (DAMP), with multiple redox states. SA suppresses both the chemoattractant activity of fully reduced HMGB1 and the increased expression of proinflammatory cytokine genes and cyclooxygenase 2 (COX-2) induced by disulfide HMGB1. Natural and synthetic SA derivatives with greater potency for inhibition of HMGB1 were identified, providing proof-of-concept that new molecules with high efficacy against sterile inflammation are attainable. An HMGB1 protein mutated in one of the SA-binding sites identified by NMR chemical shift perturbation studies retained chemoattractant activity, but lost binding of and inhibition by SA and its derivatives, thereby firmly establishing that SA binding to HMGB1 directly suppresses its proinflammatory activities. Identification of HMGB1 as a pharmacological target of SA/aspirin provides new insights into the mechanisms of action of one of the world's longest and most used natural and synthetic drugs. It may also provide an explanation for the protective effects of low-dose aspirin usage. PMID:26101955

  10. Curcumin Differs from Tetrahydrocurcumin for Molecular Targets, Signaling Pathways and Cellular Responses

    Directory of Open Access Journals (Sweden)

    Bharat B. Aggarwal

    2014-12-01

    Full Text Available Curcumin (diferuloylmethane, a golden pigment from turmeric, has been linked with antioxidant, anti-inflammatory, anticancer, antiviral, antibacterial, and antidiabetic properties. Most of the these activities have been assigned to methoxy, hydroxyl, α,β-unsaturated carbonyl moiety or to diketone groups present in curcumin. One of the major metabolites of curcumin is tetrahydrocurcumin (THC, which lacks α,β-unsaturated carbonyl moiety and is white in color. Whether THC is superior to curcumin on a molecular level is unclear and thus is the focus of this review. Various studies suggest that curcumin is a more potent antioxidant than THC; curcumin (but not THC can bind and inhibit numerous targets including DNA (cytosine-5-methyltransferase-1, heme oxygenase-1, Nrf2, β-catenin, cyclooxygenase-2, NF-kappaB, inducible nitric oxide synthase, nitric oxide, amyloid plaques, reactive oxygen species, vascular endothelial growth factor, cyclin D1, glutathione, P300/CBP, 5-lipoxygenase, cytosolic phospholipase A2, prostaglandin E2, inhibitor of NF-kappaB kinase-1, -2, P38MAPK, p-Tau, tumor necrosis factor-α, forkhead box O3a, CRAC; curcumin can inhibit tumor cell growth and suppress cellular entry of viruses such as influenza A virus and hepatitis C virus much more effectively than THC; curcumin affects membrane mobility; and curcumin is also more effective than THC in suppressing phorbol-ester-induced tumor promotion. Other studies, however, suggest that THC is superior to curcumin for induction of GSH peroxidase, glutathione-S-transferase, NADPH: quinone reductase, and quenching of free radicals. Most studies have indicated that THC exhibits higher antioxidant activity, but curcumin exhibits both pro-oxidant and antioxidant properties.

  11. Bananas, pesticides and health in southwestern Ecuador: A scalar narrative approach to targeting public health responses.

    Science.gov (United States)

    Brisbois, Benjamin

    2016-02-01

    Public health responses to agricultural pesticide exposure are often informed by ethnographic or other qualitative studies of pesticide risk perception. In addition to highlighting the importance of structural determinants of exposure, such studies can identify the specific scales at which pesticide-exposed individuals locate responsibility for their health issues, with implications for study and intervention design. In this study, an ethnographic approach was employed to map scalar features within explanatory narratives of pesticides and health in Ecuador's banana-producing El Oro province. Unstructured observation, 14 key informant interviews and 15 in-depth semi-structured interviews were carried out during 8 months of fieldwork in 2011-2013. Analysis of interview data was informed by human geographic literature on the social construction of scale. Individual-focused narratives of some participants highlighted characteristics such as carelessness and ignorance, leading to suggestions for educational interventions. More structural explanations invoked farm-scale processes, such as uncontrolled aerial fumigations on plantations owned by elites. Organization into cooperatives helped to protect small-scale farmers from 'deadly' banana markets, which in turn were linked to the Ecuadorian nation-state and actors in the banana-consuming world. These scalar elements interacted in complex ways that appear linked to social class, as more well-off individuals frequently attributed the health problems of other (poorer) people to individual behaviours, while providing more structural explanations of their own difficulties. Such individualizing narratives may help to stabilize inequitable social structures. Research implications of this study include the possibility of using scale-focused qualitative research to generate theory and candidate levels for multi-level models. Equity implications include a need for public health researchers planning interventions to engage with

  12. Enzyme responsive mesoporous silica nanoparticles for targeted tumor therapy in vitro and in vivo

    Science.gov (United States)

    Liu, Junjie; Zhang, Beilu; Luo, Zhong; Ding, Xingwei; Li, Jinghua; Dai, Liangliang; Zhou, Jun; Zhao, Xiaojing; Ye, Jingya; Cai, Kaiyong

    2015-02-01

    This study reports a biocompatible controlled drug release system based on mesoporous silica nanoparticles (MSNs) for tumor microenvironment responsive drug delivery. It was fabricated by grafting phenylboronic acid conjugated human serum albumin (PBA-HSA) onto the surfaces of MSNs as a sealing agent, via an intermediate linker of a functional polypeptide, which was composed of two functional units: the polycation cell penetrating peptide (CPP) polyarginine, and matrix metalloproteinase 2 (MMP-2) substrate peptide. A series of characterizations confirmed that the system had been successfully constructed. In vitro tests proved that the anticancer drug loading system could efficiently induce cell apoptosis in vitro. More importantly, the in vivo tumor experiments confirmed that the anticancer loading system could efficiently inhibit tumor growth with minimal side effects.This study reports a biocompatible controlled drug release system based on mesoporous silica nanoparticles (MSNs) for tumor microenvironment responsive drug delivery. It was fabricated by grafting phenylboronic acid conjugated human serum albumin (PBA-HSA) onto the surfaces of MSNs as a sealing agent, via an intermediate linker of a functional polypeptide, which was composed of two functional units: the polycation cell penetrating peptide (CPP) polyarginine, and matrix metalloproteinase 2 (MMP-2) substrate peptide. A series of characterizations confirmed that the system had been successfully constructed. In vitro tests proved that the anticancer drug loading system could efficiently induce cell apoptosis in vitro. More importantly, the in vivo tumor experiments confirmed that the anticancer loading system could efficiently inhibit tumor growth with minimal side effects. Electronic supplementary information (ESI) available: FTIR spectra, TGA curves, BET and BJH parameters, zeta potentials of nanoparticles; cleavage assay of the peptide detected by HPLC and MS; dose-dependent cytotoxicity of MSNs

  13. MicroRNA-155 targets SMAD2 and modulates the response of macrophages to transforming growth factor-{beta}.

    Science.gov (United States)

    Louafi, Fethi; Martinez-Nunez, Rocio T; Sanchez-Elsner, Tilman

    2010-12-31

    Transforming growth factor-beta (TGF-β) is a pleiotropic cytokine with important effects on processes such as fibrosis, angiogenesis, and immunosupression. Using bioinformatics, we identified SMAD2, one of the mediators of TGF-β signaling, as a predicted target for a microRNA, microRNA-155 (miR-155). MicroRNAs are a class of small non-coding RNAs that have emerged as an important class of gene expression regulators. miR-155 has been found to be involved in the regulation of the immune response in myeloid cells. Here, we provide direct evidence of binding of miR-155 to a predicted binding site and the ability of miR-155 to repress SMAD2 protein expression. We employed a lentivirally transduced monocyte cell line (THP1-155) containing an inducible miR-155 transgene to show that endogenous levels of SMAD2 protein were decreased after sustained overexpression of miR-155. This decrease in SMAD2 led to a reduction in both TGF-β-induced SMAD-2 phosphorylation and SMAD-2-dependent activation of the expression of the CAGA(12)LUC reporter plasmid. Overexpression of miR-155 altered the cellular responses to TGF-β by changing the expression of a set of genes that is involved in inflammation, fibrosis, and angiogenesis. Our study provides firm evidence of a role for miR-155 in directly repressing SMAD2 expression, and our results demonstrate the relevance of one of the two predicted target sites in SMAD2 3'-UTR. Altogether, our data uncover an important role for miR-155 in modulating the cellular response to TGF-β with possible implications in several human diseases where homeostasis of TGF-β might be altered. PMID:21036908

  14. Prostate cancer characteristics associated with response to pre-receptor targeting of the androgen axis.

    Directory of Open Access Journals (Sweden)

    Elahe A Mostaghel

    Full Text Available Factors influencing differential responses of prostate tumors to androgen receptor (AR axis-directed therapeutics are poorly understood, and predictors of treatment efficacy are needed. We hypothesized that the efficacy of inhibiting DHT ligand synthesis would associate with intra-tumoral androgen ratios indicative of relative dependence on DHT-mediated growth.We characterized two androgen-sensitive prostate cancer xenograft models after androgen suppression by castration in combination with the SRD5A inhibitor, dutasteride, as well as a panel of castration resistant metastases obtained via rapid autopsy.In LuCaP35 tumors (intra-tumoral T:DHT ratio 2:1 dutasteride suppressed DHT to 0.02 ng/gm and prolonged survival vs. castration alone (337 vs.152 days, HR 2.8, p = 0.0015. In LuCaP96 tumors (T:DHT 10:1, survival was not improved despite similar DHT reduction (0.02 ng/gm. LuCaP35 demonstrated higher expression of steroid biosynthetic enzymes maintaining DHT levels (5-fold higher SRD5A1, 41 fold higher, 99-fold higher RL-HSD, p<0.0001 for both, reconstitution of intra-tumoral DHT (to ∼30% of untreated tumors, and ∼2 fold increased expression of full length AR. In contrast, LuCaP96 demonstrated higher levels of steroid catabolizing enzymes (6.9-fold higher AKR1C2, 3000-fold higher UGT2B15, p = 0.002 and p<0.0001 respectively, persistent suppression of intra-tumoral DHT, and 6-8 fold induction of full length AR and the ligand independent V7 AR splice variant. Human metastases demonstrated bio-active androgen levels and AR full length and AR splice-variant expression consistent with the range observed in xenografts.Intrinsic differences in basal steroidogenesis, as well as variable expression of full length and splice-variant AR, associate with response and resistance to pre-receptor AR ligand suppression. Expression of steroidogenic enzymes and AR isoforms may serve as potential biomarkers of sensitivity to potent AR-axis inhibition and

  15. Measurements and simulations of the BLM response to a radiation field inside the CERF target area

    CERN Document Server

    Lebbos, E; Dehning, B; Effinger, E; Ferrari, A; Kramer, D; Nordt, A; Roeed, K; Roesler, S; Sapinski, M; Vlachoudis, V

    2010-01-01

    The CERN-EU high-energy reference field (CERF) facility is installed in one of the secondary beam lines (H6) of the Super Proton Synchrotron (SPS), in the North Experimental Area at CERN. This facility is used as a reference for testing, inter-comparing and calibrating passive and active instruments. In May 2009, the SPS provided a mixed hadron beam (protons, pions and kaons) during a few days, in order to perform several measurements with different devices such as the Radiation Protection Monitor used for residual dose rates due to Induced Radioactivity in the LHC (PMI), the Secondary Emission Monitor used for high beam losses (SEM), the Radiation Monitor for electronics (RadMon), and the Beam Loss Monitor for the LHC (BLM). This report focuses on the measurements of the BLM response during this year’s operation at CERF. The measurements evaluate the sensitivity of the BLM signal to the particle energy spectrum, with special attention to the contribution coming from thermal neutrons. For this purpose, meas...

  16. Using Complementary and Alternative Medicines to Target the Host Response during Severe Influenza

    Directory of Open Access Journals (Sweden)

    Lisa M. Alleva

    2010-01-01

    Full Text Available It is now accepted that an overwhelming inflammatory response is the cause of human deaths from avian H5N1 influenza infection. With this in mind we sought to examine the literature for examples of complementary and alternative medicines that reduce inflammation, and to place the results of this search in the context of our own work in a mouse model of influenza disease, using a pharmaceutical agent with anti-inflammatory properties. Two Chinese herbs, Angelica sinensis (Dang Gui and Salvia miltiorrhiza (Danshen, have been recently shown to protect mice during lethal experimental sepsis via inhibition of the novel inflammatory cytokine High Mobility Group Box 1 protein (HMGB1. Biochanin A, a ligand of the peroxisome proliferator activated receptors (PPAR alpha and gamma and the active isoflavone in Trifolium pratense (red clover, has anti-inflammatory properties, and thus could be used as an influenza treatment. This is of great interest since we have recently shown that gemfibrozil, a drug used to treat hyperlipidemia in humans and a synthetic ligand of PPAR alpha, significantly reduces the mortality associated with influenza infections in mice. The inflammation-modulating abilities of these natural agents should be considered in light of what is now known about the mechanisms of fatal influenza, and tested as potential candidates for influenza treatments in their own right, or as adjunct treatments to antivirals.

  17. A novel NGR-conjugated peptide targets DNA damage responses for radiosensitization.

    Science.gov (United States)

    Ma, Jinlu; Zhang, Dan; Ying, Xia; Zhao, Ying; He, Chenchen; Zhu, Qing; Han, Suxia

    2015-01-01

    Radiotherapy is one of the important treatment strategies for patients with advanced hepatocellular carcinomas. Developing novel sensitizers for radiotherapy is a key issue due to the low intrinsic radiosensitivity of hepatocellular carcinomas. It was reported the wild-type NBS1 inhibitory peptide (wtNIP) can increase radiosensitivity in several cancer cell lines by abrogating ATM-NBS1 interaction and interrupting cellular DNA damage response. Here, we developed a novel NGRconjugated peptide (NGR-sR9-wtNIP) through coupling the CNGRC angiogenic vessel-homing peptide NGR with the wtNIP peptide. Fusion peptide was tested for internalization, cytotoxicity in Hep3B cells and for tumor localization, and for toxicity in nude mice bearing human hepatocellular carcinomas xenografts. The radiosensitizing activity of NGR-sR9-wtNIP was investigated as well. We found that NGR-sR9-wtNIP can inhibit irradiation induced NBS1 phosphorylation and induce radiosensitization in Hep3B cells. When combined with IR, NGR-sR9-wtNIP suppressed tumor growth obviously in xenograft mice. In addition, the fusion peptide localized in tumor tissue specifically and barely led to any side effects on mice. Taken together, our data strongly suggest that NGRsR9- wtNIP has radiosensitizing potential for radiotherapy of hepatocellular carcinomas.

  18. Inhibition of glutathione synthesis eliminates the adaptive response of ascitic hepatoma 22 cells to nedaplatin that targets thioredoxin reductase

    Energy Technology Data Exchange (ETDEWEB)

    Wang, Yijun [School of Tea and Food Science, Anhui Agricultural University, Hefei 230036, Anhui (China); Lu, Hongjuan [Productivity Center of Jiangsu Province, Nanjing 210042, Jiangsu (China); Wang, Dongxu; Li, Shengrong; Sun, Kang; Wan, Xiaochun [School of Tea and Food Science, Anhui Agricultural University, Hefei 230036, Anhui (China); Taylor, Ethan Will [Department of Nanoscience, Joint School of Nanoscience and Nanoengineering, University of North Carolina at Greensboro, Greensboro, NC 27402 (United States); Zhang, Jinsong, E-mail: zjs@ahau.edu.cn [School of Tea and Food Science, Anhui Agricultural University, Hefei 230036, Anhui (China)

    2012-12-15

    Thioredoxin reductase (TrxR) is a target for cancer therapy and the anticancer mechanism of cisplatin involves TrxR inhibition. We hypothesize that the anticancer drug nedaplatin (NDP), an analogue of cisplatin and a second-generation platinum complex, also targets TrxR. Furthermore, we investigate whether the therapeutic efficacy of NDP can be enhanced by simultaneous modulation of 1) TrxR, via NDP, and 2) glutathione (GSH), via the GSH synthesis inhibitor buthionine sulfoximine (BSO). Mice bearing ascitic hepatoma 22 (H22) cells were treated with NDP alone or NDP plus BSO. TrxR activity of H22 cells was inhibited by NDP in a dose-dependent manner. A high correlation between the inhibition of TrxR activity at 6 h and the inhibition of ascitic fluid volume at 72 h was established (r = 0.978, p < 0.01). As an adaptive response, the viable ascitic cancer cells after NDP treatment displayed an enlarged cell phenotype, assembled with several-fold more antioxidant enzymes and GSH-predominant non-protein free thiols. This adaptive response was largely eliminated when BSO was co-administered with NDP, leading to the decimation of the H22 cell population without enhancing renal toxicity, since at this dose, NDP did not inhibit renal TrxR activity. In conclusion, the pharmacological effect of NDP involves TrxR inhibition, and the adaptive response of NDP-treated ascitic H22 cells can be efficiently counteracted by BSO. Simultaneous modulation of TrxR and GSH on ascitic H22 cells using NDP plus BSO greatly enhances therapeutic efficacy as compared with the single modulation of TrxR using NDP alone. -- Highlights: ► Nedaplatin at a pharmacological dose inhibits TrxR in cancer cells but not in kidney. ► The nedaplatin-treated cancer cells exhibit adaptive response. ► Buthionine sulfoximine inhibits glutathione in both cancer cells and kidney. ► Buthionine sulfoximine counteracts the adaptive response to the nedaplatin treatment. ► Buthionine sulfoximine does not

  19. Ordinary differential equations in affine geometry

    Directory of Open Access Journals (Sweden)

    Salvador Gigena

    1996-05-01

    Full Text Available The method of qualitative analysis is used, as applied to a class of fourth order, nonlinear ordinary differential equations, in order to classify, both locally and globally, two classes of hypersurfaces of decomposable type in affine geometry: those with constant unimodular affine mean curvature L , and those with constant Riemannian scalar curvature R. This allows to provide a large number of new examples of hypersurfaces in affine geometry.

  20. Ordinary differential equations in affine geometry

    OpenAIRE

    Salvador Gigena

    1996-01-01

    The method of qualitative analysis is used, as applied to a class of fourth order, nonlinear ordinary differential equations, in order to classify, both locally and globally, two classes of hypersurfaces of decomposable type in affine geometry: those with constant unimodular affine mean curvature L , and those with constant Riemannian scalar curvature R. This allows to provide a large number of new examples of hypersurfaces in affine geometry.

  1. Target or barrier? The cell wall of early- and later- diverging plants vs cadmium toxicity: differences in the response mechanisms

    Directory of Open Access Journals (Sweden)

    Luigi eParrotta

    2015-03-01

    Full Text Available Increasing industrialization and urbanization result in emission of pollutants in the environment including toxic heavy metals, as cadmium and lead. Among the different heavy metals contaminating the environment, cadmium raises great concern, as it is ecotoxic and as such can heavily impact ecosystems. The cell wall is the first structure of plant cells to come in contact with heavy metals. Its composition, characterized by proteins, polysaccharides and in some instances lignin and other phenolic compounds, confers the ability to bind non-covalently and/or covalently heavy metals via functional groups. A strong body of evidence in the literature has shown the role of the cell wall in heavy metal response: it sequesters heavy metals, but at the same time its synthesis and composition can be severely affected. The present review analyzes the dual property of plant cell walls, i.e. barrier and target of heavy metals, by taking Cd toxicity as example. Following a summary of the known physiological and biochemical responses of plants to Cd, the review compares the wall-related mechanisms in early- and later-diverging land plants, by considering the diversity in cell wall composition. By doing so, common as well as unique response mechanisms to metal/cadmium toxicity are identified among plant phyla and discussed. After discussing the role of hyperaccumulators’ cell walls as a particular case, the review concludes by considering important aspects for plant engineering.

  2. Ezrin, maspin, peroxiredoxin 2, and heat shock protein 27: potential targets of a streptococcal-induced autoimmune response in psoriasis.

    Science.gov (United States)

    Besgen, Petra; Trommler, Paul; Vollmer, Sigrid; Prinz, Joerg Christoph

    2010-05-01

    Psoriasis is an HLA-Cw6-associated T cell-mediated autoimmune disease of the skin that is often triggered by streptococcal angina. To identify keratinocyte proteins, which may become psoriatic autoantigens as the result of an immune response against streptococci, rabbits were immunized with heat-killed Streptococcus pyogenes. Streptococcal immunization induced Ab formation against various human keratinocyte proteins. Sera from psoriasis patients reacted against several of these proteins as well. Common serologic reactivities of rabbits and patients included the proteins ezrin, maspin, peroxiredoxin 2 (PRDX2), heat shock protein (hsp)27, and keratin 6. When used for stimulation of blood lymphocytes, ezrin, maspin, PRDX2, and hsp27 induced increased T cell activation in psoriasis patients, which was particularly evident for HLA-Cw6(+) individuals. Ag-specific T cell lines generated with these proteins consisted predominantly of CD8(+) T cells and used TCR beta-chain rearrangements, which were highly homologous to those expanded within the corresponding skin lesion. Several immunodominant epitopes on the different proteins could be defined according to sequence alignments with the whole genome of S. pyogenes. Our data indicate that maspin, ezrin, PRDX2, hsp27, and potentially keratin 6 could act as autoantigens of a streptococcal-induced autoimmune response and represent targets of the exaggerated T cell response in psoriasis. Additionally, ezrin and hsp27 might constitute antigenic links between psoriasis and inflammatory bowel disease, uveitis, or arteriosclerosis, which are clinically associated.

  3. Antibody responses to NY-ESO-1 in primary breast cancer identify a subtype target for immunotherapy.

    Science.gov (United States)

    Hamaï, Ahmed; Duperrier-Amouriaux, Karine; Pignon, Pascale; Raimbaud, Isabelle; Memeo, Lorenzo; Colarossi, Cristina; Canzonieri, Vincenzo; Perin, Tiziana; Classe, Jean-Marc; Campone, Mario; Jézéquel, Pascal; Campion, Loïc; Ayyoub, Maha; Valmori, Danila

    2011-01-01

    The highly immunogenic human tumor antigen NY-ESO-1 (ESO) is a target of choice for anti-cancer immune therapy. In this study, we assessed spontaneous antibody (Ab) responses to ESO in a large cohort of patients with primary breast cancer (BC) and addressed the correlation between the presence of anti-ESO Ab, the expression of ESO in the tumors and their characteristics. We found detectable Ab responses to ESO in 1% of the patients. Tumors from patients with circulating Ab to ESO exhibited common characteristics, being mainly hormone receptor (HR)⁻ invasive ductal carcinomas of high grade, including both HER2⁻ and HER2⁺ tumors. In line with these results, we detected ESO expression in 20% of primary HR⁻ BC, including both ESO Ab⁺ and Ab⁻ patients, but not in HR⁺ BC. Interestingly, whereas expression levels in ESO⁺ BC were not significantly different between ESO Ab⁺ and Ab⁻ patients, the former had, in average, significantly higher numbers of tumor-infiltrated lymph nodes, indicating that lymph node invasion may be required for the development of spontaneous anti-tumor immune responses. Thus, the presence of ESO Ab identifies a tumor subtype of HR⁻ (HER2⁻ or HER2⁺) primary BC with frequent ESO expression and, together with the assessment of antigen expression in the tumor, may be instrumental for the selection of patients for whom ESO-based immunotherapy may complement standard therapy.

  4. Metabolomics reveals metabolic targets and biphasic responses in breast cancer cells treated by curcumin alone and in association with docetaxel.

    Directory of Open Access Journals (Sweden)

    Mathilde Bayet-Robert

    Full Text Available BACKGROUND: Curcumin (CUR has deserved extensive research due to its anti-inflammatory properties, of interest in human diseases including cancer. However, pleiotropic even paradoxical responses of tumor cells have been reported, and the mechanisms of action of CUR remain uncompletely elucidated. METHODOLOGY/PRINCIPAL FINDINGS: (1H-NMR spectroscopy-based metabolomics was applied to get novel insight into responses of MCF7 and MDA-MB-231 breast cancer cells to CUR alone, and MCF7 cells to CUR in cotreatment with docetaxel (DTX. In both cell types, a major target of CUR was glutathione metabolism. Total glutathione (GSx increased at low dose CUR (≤ 10 mg.l(-1-28 µM- (up to +121% in MCF7 cells, P<0.01, and +138% in MDA-MB-231 cells, P<0.01, but decreased at high dose (≥ 25 mg.l(-1 -70 µM- (-49%, in MCF7 cells, P<0.02, and -56% in MDA-MB-231 cells, P<0.025. At high dose, in both cell types, GSx-related metabolites decreased, including homocystein, creatine and taurine (-60 to -80%, all, P<0.05. Together with glutathione-S-transferase actvity, data established that GSx biosynthesis was upregulated at low dose, and GSx consumption activated at high dose. Another major target, in both cell types, was lipid metabolism involving, at high doses, accumulation of polyunsaturated and total free fatty acids (between ×4.5 and ×11, P<0.025, and decrease of glycerophospho-ethanolamine and -choline (about -60%, P<0.025. Multivariate statistical analyses showed a metabolic transition, even a biphasic behavior of some metabolites including GSx, between low and high doses. In addition, CUR at 10 mg.l(-1 in cotreatment with DTX induced modifications in glutathione metabolism, lipid metabolism, and glucose utilization. Some of these changes were biphasic depending on the duration of exposure to CUR. CONCLUSIONS/SIGNIFICANCE: Metabolomics reveals major metabolic targets of CUR in breast cancer cells, and biphasic responses that challenge the widely accepted

  5. Affine connections, midpoint formation, and point reflection

    DEFF Research Database (Denmark)

    Kock, Anders

    2011-01-01

    We describe some differential-geometric structures in combinatorial terms: namely affine connections and their torsion and curvature, and we show that torsion free affine connections may equivalently be presented in terms of some simpler combinatorial structure: midpoint formation, and point refl...... reflection (geodesic symmetry). The method employed is that of synthetic differential geometry, which is briefly explained.......We describe some differential-geometric structures in combinatorial terms: namely affine connections and their torsion and curvature, and we show that torsion free affine connections may equivalently be presented in terms of some simpler combinatorial structure: midpoint formation, and point...

  6. A Novel Vertex Affinity for Community Detection

    Energy Technology Data Exchange (ETDEWEB)

    Yoo, Andy [Lawrence Livermore National Lab. (LLNL), Livermore, CA (United States); Sanders, Geoffrey [Lawrence Livermore National Lab. (LLNL), Livermore, CA (United States); Henson, Van [Lawrence Livermore National Lab. (LLNL), Livermore, CA (United States); Vassilevski, Panayot [Lawrence Livermore National Lab. (LLNL), Livermore, CA (United States)

    2015-10-05

    We propose a novel vertex affinity measure in this paper. The new vertex affinity quantifies the proximity between two vertices in terms of their clustering strength and is ideal for such graph analytics applications as community detection. We also developed a framework that combines simple graph searches and resistance circuit formulas to compute the vertex affinity efficiently. We study the properties of the new affinity measure empirically in comparison to those of other popular vertex proximity metrics. Our results show that the existing metrics are ill-suited for community detection due to their lack of fundamental properties that are essential for correctly capturing inter- and intra-cluster vertex proximity.

  7. Targeted demersal fish species exhibit variable responses to long-term protection from fishing at the Houtman Abrolhos Islands

    Science.gov (United States)

    Bornt, Katrina R.; McLean, Dianne L.; Langlois, Tim J.; Harvey, Euan S.; Bellchambers, Lynda M.; Evans, Scott N.; Newman, Stephen J.

    2015-12-01

    Natural fluctuations in the abundance and length of targeted fish are often disrupted by acute environmental changes and anthropogenic impacts, particularly fishing pressure. Long-term assessments of targeted fish populations inside and outside areas closed to fishing are often necessary to elucidate these effects, yet few of these studies extend over long time periods. We assessed trends in the abundance and length of six targeted fish species in areas open and closed to fishing on seven occasions spanning a 9-year period (2005-2010 and 2013) at the Houtman Abrolhos Islands, Western Australia. Shallow (8-12 m) and deep (22-26 m) coral-dominated reef sites were sampled across four geographically separated island groups using baited remote underwater stereo-video (stereo-BRUV). Between 2005 and 2010, populations of Lethrinus miniatus, Lethrinus nebulosus, Plectropomus leopardus, and Chrysophrys auratus became increasingly dominated by larger individuals, potentially indicative of an ageing population. Between 2010 and 2013, however, there was a significant increase in the proportion of smaller L. miniatus, L. nebulosus, and P. leopardus in both open and closed areas, reflecting increased recruitment perhaps due to changing environmental conditions associated with a marine heat wave anomaly. This recruitment pulse was not observed for the other species in this study ( Chr. auratus, Choerodon rubescens, and Glaucosoma hebraicum). Lethrinus miniatus, L. nebulosus, Chr. auratus, and P. leopardus were larger in closed areas relative to open areas; however, they were not more abundant. These complex responses to protection also varied across sampling years for certain species (e.g., P. leopardus). Monitoring changes over the long-term in areas open and closed to fishing provides a sound basis for separating environmental variability from that associated with fishing mortality, which is crucial for optimising fisheries management.

  8. Global connectivity of hub residues in Oncoprotein structures encodes genetic factors dictating personalized drug response to targeted Cancer therapy

    Science.gov (United States)

    Soundararajan, Venky; Aravamudan, Murali

    2014-12-01

    The efficacy and mechanisms of therapeutic action are largely described by atomic bonds and interactions local to drug binding sites. Here we introduce global connectivity analysis as a high-throughput computational assay of therapeutic action - inspired by the Google page rank algorithm that unearths most ``globally connected'' websites from the information-dense world wide web (WWW). We execute short timescale (30 ps) molecular dynamics simulations with high sampling frequency (0.01 ps), to identify amino acid residue hubs whose global connectivity dynamics are characteristic of the ligand or mutation associated with the target protein. We find that unexpected allosteric hubs - up to 20Å from the ATP binding site, but within 5Å of the phosphorylation site - encode the Gibbs free energy of inhibition (ΔGinhibition) for select protein kinase-targeted cancer therapeutics. We further find that clinically relevant somatic cancer mutations implicated in both drug resistance and personalized drug sensitivity can be predicted in a high-throughput fashion. Our results establish global connectivity analysis as a potent assay of protein functional modulation. This sets the stage for unearthing disease-causal exome mutations and motivates forecast of clinical drug response on a patient-by-patient basis. We suggest incorporation of structure-guided genetic inference assays into pharmaceutical and healthcare Oncology workflows.

  9. pH-Responsive Hyaluronic Acid-Based Mixed Micelles for the Hepatoma-Targeting Delivery of Doxorubicin

    Directory of Open Access Journals (Sweden)

    Jing-Liang Wu

    2016-03-01

    Full Text Available The tumor targetability and stimulus responsivity of drug delivery systems are crucial in cancer diagnosis and treatment. In this study, hepatoma-targeting mixed micelles composed of a hyaluronic acid–glycyrrhetinic acid conjugate and a hyaluronic acid-l-histidine conjugate (HA–GA/HA–His were prepared through ultrasonic dispersion. The formation and characterization of the mixed micelles were confirmed via 1H-NMR, particle size, and ζ potential measurements. The in vitro cellular uptake of the micelles was evaluated using human liver carcinoma (HepG2 cells. The antitumor effect of doxorubicin (DOX-loaded micelles was investigated in vitro and in vivo. Results indicated that the DOX-loaded HA–GA/HA–His micelles showed a pH-dependent controlled release and were remarkably absorbed by HepG2 cells. Compared with free DOX, the DOX-loaded HA–GA/HA–His micelles showed a higher cytotoxicity to HepG2 cells. Moreover, the micelles effectively inhibited tumor growth in H22 cell-bearing mice. These results suggest that the HA–GA/HA–His mixed micelles are a good candidate for drug delivery in the prevention and treatment of hepatocarcinoma.

  10. Analyses of Potential Predictive Markers and Response to Targeted Therapy in Patients with Advanced Clear-cell Renal Cell Carcinoma

    Directory of Open Access Journals (Sweden)

    Yan Song

    2015-01-01

    Full Text Available Background: Vascular endothelial growth factor-targeted agents are standard treatments in advanced clear-cell renal cell carcinoma (ccRCC, but biomarkers of activity are lacking. The aim of this study was to investigate the association of Von Hippel-Lindau (VHL gene status, vascular endothelial growth factor receptor (VEGFR or stem cell factor receptor (KIT expression, and their relationships with characteristics and clinical outcome of advanced ccRCC. Methods: A total of 59 patients who received targeted treatment with sunitinib or pazopanib were evaluated for determination at Cancer Hospital and Institute, Chinese Academy of Medical Sciences between January 2010 and November 2012. Paraffin-embedded tumor samples were collected and status of the VHL gene and expression of VEGFR and KIT were determined by VHL sequence analysis and immunohistochemistry. Clinical-pathological features were collected and efficacy such as response rate and Median progression-free survival (PFS and overall survival (OS were calculated and then compared based on expression status. The Chi-square test, the Kaplan-Meier method, and the Lon-rank test were used for statistical analyses. Results: Of 59 patients, objective responses were observed in 28 patients (47.5%. The median PFS was 13.8 months and median OS was 39.9 months. There was an improved PFS in patients with the following clinical features: Male gender, number of metastatic sites 2 or less, VEGFR-2 positive or KIT positive. Eleven patients (18.6% had evidence of VHL mutation, with an objective response rate of 45.5%, which showed no difference with patients with no VHL mutation (47.9%. VHL mutation status did not correlate with either overall response rate (P = 0.938 or PFS (P = 0.277. The PFS was 17.6 months and 22.2 months in VEGFR-2 positive patients and KIT positive patients, respectively, which was significantly longer than that of VEGFR-2 or KIT negative patients (P = 0.026 and P = 0.043. Conclusion

  11. Analyses of Potential Predictive Markers and Response to Targeted Therapy in Patients with Advanced Clear-cell Renal Cell Carcinoma

    Institute of Scientific and Technical Information of China (English)

    Yan Song; Jing Huang; Ling Shan; Hong-Tu Zhang

    2015-01-01

    Background:Vascular endothelial growth factor-targeted agents are standard treatments in advanced clear-cell renal cell carcinoma (ccRCC),but biomarkers of activity are lacking.The aim of this study was to investigate the association of Von Hippel-Lindau (VHL) gene status,vascular endothelial growth factor receptor (VEGFR) or stem cell factor receptor (KIT) expression,and their relationships with characteristics and clinical outcome of advanced ccRCC.Methods:A total of 59 patients who received targeted treatment with sunitinib or pazopanib were evaluated for determination at Cancer Hospital and Institute,Chinese Academy of Medical Sciences between January 2010 and November 2012.Paraffin-embedded tumor samples were collected and status of the VHL gene and expression of VEGFR and KIT were determined by VHL sequence analysis and immunohistochemistry.Clinical-pathological features were collected and efficacy such as response rate and Median progression-free survival (PFS) and ovcrall survival (OS) were calculated and then compared based on expression status.The Chi-square test,the KaplanMeier method,and the Lon-rank test were used for statistical analyses.Results:Of 59 patients,objective responses were observed in 28 patients (47.5%).The median PFS was 13.8 months and median OS was 39.9 months.There was an improved PFS in patients with the following clinical features:Male gender,number of metastatic sites 2 or less,VEGFR-2 positive or KIT positive.Eleven patients (18.6%) had evidence of VHL mutation,with an objective response rate of 45.5%,which showed no difference with patients with no VHL mutation (47.9%).VHL mutation status did not correlate with either overall response rate (P =0.938) or PFS (P =0.277).The PFS was 17.6 months and 22.2 months in VEGFR-2 positive patients and KIT positive patients,respectively,which was significantly longer than that of VEGFR-2 or KIT negative patients (P =0.026 and P =0.043).Conclusion:VHL mutation status could not predict

  12. Influence of the APOE genotype on hepatic stress response: Studies in APOE targeted replacement mice and human liver cells.

    Science.gov (United States)

    Dose, Janina; Nebel, Almut; Piegholdt, Stefanie; Rimbach, Gerald; Huebbe, Patricia

    2016-07-01

    Apolipoprotein E (APOE) is a multifunctional plasma protein mainly acting in lipid metabolism. Human APOE is polymorphic with three major isoforms (APOE2, APOE3 and APOE4). Up to 75% of the body's APOE is produced by the liver. There is increasing evidence from studies in brain-derived cells that APOE4 affects mitochondrial function and biogenesis as well as stress and inflammatory responses - processes, whose disturbances are considered hallmarks of the ageing process. However, although the liver is the main production site of APOE, knowledge about the impact of the APOE genotype on hepatic stress response-related processes is rather limited. Therefore, we studied biomarkers of oxidative status (glutathione levels, 3-nitrotyrosine adducts, protein carbonyl concentration), ER stress (XBP1(S), BiP, DDIT3), proteasome activity, mitochondrial function (respiratory complexes, ATP levels and mitochondrial membrane potential as well as biomarkers of mitochondrial biogenesis, fission and fusion), autophagy (LC3, LAMP2A), apoptosis (BCL2, BAX, CYCS) and DNA damage in the liver of APOE targeted replacement (TR) mice and in Huh7 hepatocytes overexpressing the APOE3 and the APOE4 isoform, respectively. APOE4 mice exhibited a lower chymotrypsin-like and a higher trypsin-like proteasome activity. Levels of protein carbonyls were moderately higher in liver tissue of APOE4 vs. APOE3 mice. Other biomarkers of oxidative stress were similar between the two genotypes. Under basal conditions, the stress-response pathways investigated appeared largely unaffected by the APOE genotype. However, upon stress induction, APOE4 expressing cells showed lower levels of adenosine triphosphate (ATP) and lower mRNA levels of the ATP-generating complex V of the mitochondrial respiratory chain. Overall, our findings provide evidence for a rather low influence of the APOE genotype on the hepatic stress response processes investigated in this study. PMID:27130033

  13. Magnetic actuated pH-responsive hydrogel-based soft micro-robot for targeted drug delivery

    Science.gov (United States)

    Li, Hao; Go, Gwangjun; Ko, Seong Yong; Park, Jong-Oh; Park, Sukho

    2016-02-01

    For drug delivery in cancer therapy, various stimuli-responsive hydrogel-based micro-devices have been studied with great interest. Here, we present a new concept for a hybrid actuated soft microrobot targeted drug delivery. The proposed soft microrobot consists of a hydrogel bilayer structure of 2-hydroxyethyl methacrylate (PHEMA) and poly (ethylene glycol) acrylate (PEGDA) with iron (II, III) oxide particles (Fe3O4). The PHEMA layer as a pH-responsive gel is used for a trapping and unfolding motion of the soft microrobot in pH-varying solution, and the PEGDA-with-Fe3O4 layer is employed for the locomotion of the soft microrobot in the magnetic field. The bilayer soft microrobot was fabricated by a conventional photolithography procedure and its characteristics were analyzed and presented. To evaluate the trapping performance and the motility of the soft microrobot, test solutions with different pH values and an electromagnetic actuation (EMA) system were used. First, the soft microrobot showed its full trapping motion at about pH 9.58 and its unfolding motion at about pH 2.6. Second, the soft microrobot showed a moving velocity of about 600 μm s-1 through the generated magnetic field of the EMA system. Finally, we fabricated the real anti-cancer drug microbeads (PCL-DTX) and executed the cytotoxicity test using the mammary carcinoma cells (4T1). The viability of the 4T1 cells treated with the proposed microrobot and the PCL-DTX microbeads decreased to 70.25 ± 1.52%. The result demonstrated that the soft microrobot can be moved to a target position by the EMA system and can release a small amount of beads by the pH variation and the robot exhibited no toxicity to the cells. In the future, we expect that the proposed soft microrobot can be applied to a new tumor-therapeutic tool that can move to a target tumor and release anti-tumor drugs.

  14. Affine processes on positive semidefinite matrices

    CERN Document Server

    Cuchiero, Christa; Mayerhofer, Eberhard; Teichmann, Josef

    2009-01-01

    This paper provides the mathematical foundation for stochastically continuous affine processes on the cone of positive semidefinite symmetric matrices. These matrix-valued affine processes have arisen from a large and growing range of useful applications in finance, including multi-asset option pricing with stochastic volatility and correlation structures, and fixed-income models with stochastically correlated risk factors and default intensities.

  15. Lectures on extended affine Lie algebras

    CERN Document Server

    Neher, Erhard

    2010-01-01

    We give an introduction to the structure theory of extended affine Lie algebras, which provide a common framework for finite-dimensional semisimple, affine and toroidal Lie algebras. The notes are based on a lecture series given during the Fields Institute summer school at the University of Ottawa in June 2009.

  16. Global affine differential geometry of hypersurfaces

    CERN Document Server

    Li, An-Min; Zhao, Guosong; Hu, Zejun

    2015-01-01

    This book draws a colorful and widespread picture of global affine hypersurface theory up to the most recent state. Moreover, the recent development revealed that affine differential geometry- as differential geometry in general- has an exciting intersection area with other fields of interest, like partial differential equations, global analysis, convex geometry and Riemann surfaces.

  17. Dyes with high affinity for polylactide

    Institute of Scientific and Technical Information of China (English)

    Liang He; Shu Fen Zhang; Bing Tao Tang; Li Li Wang; Jin Zong Yang

    2007-01-01

    Attempts were made to develop dyes with high affinity for polylactide as an alternative to the existent commercial disperse dyes.The dyes synthesized according to the affinity concept of dye to polylactide exhibited excellent dyeing properties on polylactide compared with the commercial disperse dyes.

  18. Free C+ actions on affine threefolds

    OpenAIRE

    Kraft, H.

    2005-01-01

    We study algebraic actions of the additive group C+ on an affine threefold X and prove a smoothness property for the quotient morphism X -< X//C+. Then, following Shulim Kaliman, we give a proof of the conjecture that every free C+ action on affine 3-space C^3 is a translation.

  19. Porosity of Self-affine Sets

    Institute of Scientific and Technical Information of China (English)

    Lifeng XI

    2008-01-01

    In this paper,it is proved that any self-affine set satisfying the strong separation condition is uniformly porous.The author constructs a self-affine set which is not porous,although the open set condition holds.Besides,the author also gives a C1 iterated function system such that its invariant set is not porous.

  20. On affine non-negative matrix factorization

    DEFF Research Database (Denmark)

    Laurberg, Hans; Hansen, Lars Kai

    2007-01-01

    We generalize the non-negative matrix factorization (NMF) generative model to incorporate an explicit offset. Multiplicative estimation algorithms are provided for the resulting sparse affine NMF model. We show that the affine model has improved uniqueness properties and leads to more accurate...

  1. Phosphopeptide enrichment by immobilized metal affinity chromatography

    DEFF Research Database (Denmark)

    Thingholm, Tine E.; Larsen, Martin R.

    2016-01-01

    Immobilized metal affinity chromatography (IMAC) has been the method of choice for phosphopeptide enrichment prior to mass spectrometric analysis for many years and it is still used extensively in many laboratories. Using the affinity of negatively charged phosphate groups towards positively...

  2. Photosensitizer enhanced disassembly of amphiphilic micelle for ROS-response targeted tumor therapy in vivo.

    Science.gov (United States)

    Dai, Liangliang; Yu, Yonglin; Luo, Zhong; Li, Menghuan; Chen, Weizhen; Shen, Xinkun; Chen, Feng; Sun, Qiang; Zhang, Qingfeng; Gu, Hao; Cai, Kaiyong

    2016-10-01

    This study reports a reactive oxygen species (ROS) sensitive drug delivery system based on amphiphilic polymer of poly(propylene sulfide)-polyethylene glycol-serine-folic acid (PPS-mPEG-Ser-FA). The polymer could form homogeneous micelles with an average diameter of around 80 nm through self-assembly, which would then be loaded with the singlet oxygen-generating photosensitizer of zinc phthalocyanine (ZNPC) and anti-cancer drug of DOX. The disassembly of micelles could be triggered by the hydrophobic to hydrophilic transition of the PPS core in response to ROS-induced oxidation in vitro. ZNPC molecules are capable of producing ROS under laser irradiation, which results in the rapid disassembly of micelles and releasing of the anti-tumor drug for tumor therapy under physiological condition otherwise. Moreover, the excessive ROS production deriving from ZNPC synergically induces cells apoptosis. Furthermore, the DOX loaded amphiphilic micelles could be internalized by tumor cells via FA receptor-mediated endocytosis to effectively inhibit the tumor growth in vivo, while with only minimal toxic side effects. The results in vitro and in vivo consistently demonstrate that the light-responsive micelle is a promising biodegradable nanocarrier for on-command drug release and targeted tumor therapy. PMID:27423095

  3. Non-Target Impacts of an Attract-and-Kill Formulation Based on Plant Volatiles: Responses of some Generalist Predators.

    Science.gov (United States)

    Gregg, Peter C; Del Socorro, Alice P; Binns, Matthew R

    2016-07-01

    Responses of non-target insects to a blend of plant volatiles used as components in an attract-and-kill formulation for Helicoverpa spp. (Lepidoptera: Noctuidae) were studied in an Australian cotton field. Two experiments, one involving suction sampling during the day and the other at night, were conducted. Rows that had been treated with the volatile blend, with no added insecticide, were sampled with a large suction sampler 18, 42, and 85 h (day experiment) and 6, 30, and 78 h (night experiment) after treatment. Rows located 5, 10, 20, and 300 m away from the treated row were similarly sampled. Of seven generalist predators, only one accumulated on the treated rows compared to the untreated rows. Of the other six, five were found in lower numbers on the treated rows, and for one no significant effects were detected. Compared to pre-spray baseline levels, numbers of several taxa increased across the whole field after spraying, suggesting area-wide attraction, but localized responses to the treated rows were weak, and apparent repellence was more common than attraction. We suggest that attract-and-kill with plant volatiles should have minimal effects on populations of these predators, and is likely to be compatible with integrated pest management.

  4. Pyruvate Carboxylase Activates the RIG-I-like Receptor-Mediated Antiviral Immune Response by Targeting the MAVS signalosome

    Science.gov (United States)

    Cao, Zhongying; Zhou, Yaqin; Zhu, Shengli; Feng, Jian; Chen, Xueyuan; Liu, Shi; Peng, Nanfang; Yang, Xiaodan; Xu, Gang; Zhu, Ying

    2016-01-01

    When retinoic acid-inducible gene 1 protein (RIG-I)-like receptors sense viral dsRNA in the cytosol, RIG-I and melanoma differentiation-associated gene 5 (MDA5) are recruited to the mitochondria to interact with mitochondrial antiviral signaling protein (MAVS) and initiate antiviral immune responses. In this study, we demonstrate that the biotin-containing enzyme pyruvate carboxylase (PC) plays an essential role in the virus-triggered activation of nuclear factor kappa B (NF-κB) signaling mediated by MAVS. PC contributes to the enhanced production of type I interferons (IFNs) and pro-inflammatory cytokines, and PC knockdown inhibits the virus-triggered innate immune response. In addition, PC shows extensive antiviral activity against RNA viruses, including influenza A virus (IAV), human enterovirus 71 (EV71), and vesicular stomatitis virus (VSV). Furthermore, PC mediates antiviral action by targeting the MAVS signalosome and induces IFNs and pro-inflammatory cytokines by promoting phosphorylation of NF-κB inhibitor-α (IκBα) and the IκB kinase (IKK) complex, as well as NF-κB nuclear translocation, which leads to activation of interferon-stimulated genes (ISGs), including double-stranded RNA-dependent protein kinase (PKR) and myxovirus resistance protein 1 (Mx1). Our findings suggest that PC is an important player in host antiviral signaling. PMID:26906558

  5. Improving image segmentation by learning region affinities

    Energy Technology Data Exchange (ETDEWEB)

    Prasad, Lakshman [Los Alamos National Laboratory; Yang, Xingwei [TEMPLE UNIV.; Latecki, Longin J [TEMPLE UNIV.

    2010-11-03

    We utilize the context information of other regions in hierarchical image segmentation to learn new regions affinities. It is well known that a single choice of quantization of an image space is highly unlikely to be a common optimal quantization level for all categories. Each level of quantization has its own benefits. Therefore, we utilize the hierarchical information among different quantizations as well as spatial proximity of their regions. The proposed affinity learning takes into account higher order relations among image regions, both local and long range relations, making it robust to instabilities and errors of the original, pairwise region affinities. Once the learnt affinities are obtained, we use a standard image segmentation algorithm to get the final segmentation. Moreover, the learnt affinities can be naturally unutilized in interactive segmentation. Experimental results on Berkeley Segmentation Dataset and MSRC Object Recognition Dataset are comparable and in some aspects better than the state-of-art methods.

  6. Reflectable bases for affine reflection systems

    CERN Document Server

    Azam, Saeid; Yousofzadeh, Malihe

    2011-01-01

    The notion of a "root base" together with its geometry plays a crucial role in the theory of finite and affine Lie theory. However, it is known that such a notion does not exist for the recent generalizations of finite and affine root systems such as extended affine root systems and affine reflection systems. As an alternative, we introduce the notion of a "reflectable base", a minimal subset $\\Pi$ of roots such that the non-isotropic part of the root system can be recovered by reflecting roots of $\\Pi$ relative to the hyperplanes determined by $\\Pi$. We give a full characterization of reflectable bases for tame irreducible affine reflection systems of reduced types, excluding types $E_{6,7,8}$. As a byproduct of our results, we show that if the root system under consideration is locally finite then any reflectable base is an integral base.

  7. Fatty acid and drug binding to a low-affinity component of human serum albumin, purified by affinity chromatography

    DEFF Research Database (Denmark)

    Vorum, H; Pedersen, A O; Honoré, B

    1992-01-01

    of two albumin components about 40% of the albumin having high affinity and about 60% having low affinity. By affinity chromatography we succeeded in purifying the low-affinity component from the mixture. The high-affinity component, however, could not be isolated. We further analyzed the fatty acid...... and drug binding abilities of the low-affinity component. The fatty acids decanoate, laurate, myristate and palmitate were bound with higher affinity to the mixture than to the low-affinity component. Diazepam was bound with nearly the same affinity to the low-affinity component as to the albumin mixture...

  8. Targeting of Ly9 (CD229) Disrupts Marginal Zone and B1 B Cell Homeostasis and Antibody Responses.

    Science.gov (United States)

    Cuenca, Marta; Romero, Xavier; Sintes, Jordi; Terhorst, Cox; Engel, Pablo

    2016-01-15

    Marginal zone (MZ) and B1 B cells have the capacity to respond to foreign Ags more rapidly than conventional B cells, providing early immune responses to blood-borne pathogens. Ly9 (CD229, SLAMF3), a member of the signaling lymphocytic activation molecule family receptors, has been implicated in the development and function of innate T lymphocytes. In this article, we provide evidence that in Ly9-deficient mice splenic transitional 1, MZ, and B1a B cells are markedly expanded, whereas development of B lymphocytes in bone marrow is unaltered. Consistent with an increased number of these B cell subsets, we detected elevated levels of IgG3 natural Abs and a striking increase of T-independent type II Abs after immunization with 2,4,6-trinitrophenyl-Ficoll in the serum of Ly9-deficient mice. The notion that Ly9 could be a negative regulator of innate-like B cell responses was supported by the observation that administering an mAb directed against Ly9 to wild-type mice selectively eliminated splenic MZ B cells and significantly reduced the numbers of B1 and transitional 1 B cells. In addition, Ly9 mAb dramatically diminished in vivo humoral responses and caused a selective downregulation of the CD19/CD21/CD81 complex on B cells and concomitantly an impaired B cell survival and activation in an Fc-independent manner. We conclude that altered signaling caused by the absence of Ly9 or induced by anti-Ly9 may negatively regulate development and function of innate-like B cells by modulating B cell activation thresholds. The results suggest that Ly9 could serve as a novel target for the treatment of B cell-related diseases.

  9. The relation of morphology and affinity maturation in germinal centers

    CERN Document Server

    Meyer-Hermann, M

    2002-01-01

    The specific morphology of germinal centers is analyzed in the context of the optimization of the humoral immune response. The relevance of dark and light zones for the affinity maturation process is investigated in the framework of a theoretical model for the germinal center reaction. Especially, it is shown that an intermediate appearance of dark zones in germinal center reactions is advantageous for the process of antibody optimization.

  10. Non-Zenoness of piecewise affine dynamical systems and affine complementarity systems with inputs

    Institute of Scientific and Technical Information of China (English)

    Le Quang THUAN

    2014-01-01

    In the context of continuous piecewise affine dynamical systems and affine complementarity systems with inputs, we study the existence of Zeno behavior, i.e., infinite number of mode transitions in a finite-length time interval, in this paper. The main result reveals that continuous piecewise affine dynamical systems with piecewise real-analytic inputs do not exhibit Zeno behavior. Applied the achieved result to affine complementarity systems with inputs, we also obtained a similar conclusion. A direct benefit of the main result is that one can apply smooth ordinary differential equations theory in a local manner for the analysis of continuous piecewise affine dynamical systems with inputs.

  11. Targeting autophagy overcomes Enzalutamide resistance in castration-resistant prostate cancer cells and improves therapeutic response in a xenograft model

    Science.gov (United States)

    Nguyen, H G; Yang, J C; Kung, H-J; Shi, X-B; Tilki, D; Lara, P N; DeVere White, R W; Gao, A C; Evans, C P

    2014-01-01

    Macro-autophagy is associated with drug resistance in various cancers and can function as an adaptive response to maintain cell survival under metabolic stresses, including androgen deprivation. Androgen deprivation or treatment with androgen receptor (AR) signaling inhibitor (ARSI), Enzalutamide (MDV-3100, ENZA) or bicalutamide induced autophagy in androgen-dependent and in castration-resistant CaP (castration-resistant prostate cancer (CRPC)) cell lines. The autophagic cascade triggered by AR blockage, correlated with the increased light chain 3-II/I ratio and ATG-5 expression. Autophagy was observed in a subpopulation of C4-2B cells that developed insensitivity to ENZA after sustained exposure in culture. Using flow cytometry and clonogenic assays, we showed that inhibiting autophagy with clomipramine (CMI), chloroquine or metformin increased apoptosis and significantly impaired cell viability. This autophagic process was mediated by AMP-dependent protein kinase (AMPK) activation and the suppression of mammalian target of rapamycin (mTOR) through Raptor phosphorylation (Serine 792). Furthermore, small interfering RNA targeting AMPK significantly inhibited autophagy and promoted cell death in CaP cells acutely or chronically exposed to ENZA or androgen deprivation, suggesting that autophagy is an important survival mechanism in CRPC. Lastly, in vivo studies with mice orthotopically implanted with ENZA-resistant cells demonstrated that the combination of ENZA and autophagy modulators, CMI or metformin significantly reduced tumor growth when compared with control groups (P<0.005). In conclusion, autophagy is as an important mechanism of resistance to ARSI in CRPC. Antiandrogen-induced autophagy is mediated through the activation of AMPK pathway and the suppression of mTOR pathway. Blocking autophagy pharmacologically or genetically significantly impairs prostate cancer cell survival in vitro and in vivo, implying the therapeutics potential of autophagy inhibitors

  12. The membrane targeted apoptosis modulators erucylphosphocholine and erucylphosphohomocholine increase the radiation response of human glioblastoma cell lines in vitro

    International Nuclear Information System (INIS)

    Alkylphosphocholines constitute a novel class of antineoplastic synthetic phospholipid derivatives that induce apoptosis of human tumor cell lines by targeting cellular membranes. We could recently show that the first intravenously applicable alkylphosphocholine erucylphosphocholine (ErPC) is a potent inducer of apoptosis in highly resistant human astrocytoma/glioblastoma cell lines in vitro. ErPC was shown to cross the blood brain barrier upon repeated intravenous injections in rats and thus constitutes a promising candidate for glioblastoma therapy. Aim of the present study was to analyze putative beneficial effects of ErPC and its clinically more advanced derivative erucylphosphohomocholine (erucyl-N, N, N-trimethylpropanolaminphosphate, ErPC3, Erufosine™ on radiation-induced apoptosis and eradication of clonogenic tumor cells in human astrocytoma/glioblastoma cell lines in vitro. While all cell lines showed high intrinsic resistance against radiation-induced apoptosis as determined by fluorescence microscopy, treatment with ErPC and ErPC3 strongly increased sensitivity of the cells to radiation-induced cell death (apoptosis and necrosis). T98G cells were most responsive to the combined treatment revealing highly synergistic effects while A172 showed mostly additive to synergistic effects, and U87MG cells sub-additive, additive or synergistic effects, depending on the respective radiation-dose, drug-concentration and treatment time. Combined treatment enhanced therapy-induced damage of the mitochondria and caspase-activation. Importantly, combined treatment also increased radiation-induced eradication of clonogenic T98G cells as determined by standard colony formation assays. Our observations make the combined treatment with ionizing radiation and the membrane targeted apoptosis modulators ErPC and ErPC3 a promising approach for the treatment of patients suffering from malignant glioma. The use of this innovative treatment concept in an in vivo xenograft

  13. Galactose targeted pH-responsive copolymer conjugated with near infrared fluorescence probe for imaging of intelligent drug delivery.

    Science.gov (United States)

    Fu, Liyi; Sun, Chunyang; Yan, Lifeng

    2015-01-28

    Theranostic polymeric nanomaterials are of special important in cancer treatment. Here, novel galactose targeted pH-responsive amphiphilic multiblock copolymer conjugated with both drug and near-infrared fluorescence (NIR) probe has been designed and prepared by a four-steps process: (1) ring-opening polymerization (ROP) of N-carboxy anhydride (NCA) monomers using propargylamine as initiator; (2) reversible addition-fragmentation chain transfer (RAFT) polymerization of oligo(ethylene glycol) methacrylate (OEGMA) and gal monomer by an azido modified RAFT agent; (3) combing the obtained two polymeric segments by click reaction; (4) NIR copolymer prodrug was synthesized by chemical linkage of both cyanine dye and anticancer drug doxorubicin to the block copolymer via amide bond and hydrazone, respectively. The obtained NIRF copolymers were characterized by nuclear magnetic resonance (NMR), gel permeation chromatography (GPC), and its was measured by means of micelles dynamic light scattering (DLS), field emission transmission electron microscopy (FETEM), and UV-vis and fluorescence spectrophotometry. The prodrug has strong fluorescence in the near-infrared region, and a pH sensitive drug release was confirmed at pH of 5.4 via an in vitro drug release experiment. Confocal laser scanning microscopy (CLSM) and flow cytometry experiments of the prodrug on both HepG2 and NIH3T3 cells reveal that the galactose targeted polymeric prodrug shows a fast and enhanced endocytosis due to the specific interaction for HepG2 cells, indicating the as-prepared polymer is a candidate for theranosis of liver cancer. PMID:25569169

  14. Cellular stress response: a novel target for chemoprevention and nutritional neuroprotection in aging, neurodegenerative disorders and longevity.

    Science.gov (United States)

    Calabrese, Vittorio; Cornelius, Carolin; Mancuso, Cesare; Pennisi, Giovanni; Calafato, Stella; Bellia, Francesco; Bates, Timothy E; Giuffrida Stella, Anna Maria; Schapira, Tony; Dinkova Kostova, Albena T; Rizzarelli, Enrico

    2008-12-01

    The predominant molecular symptom of aging is the accumulation of altered gene products. Moreover, several conditions including protein, lipid or glucose oxidation disrupt redox homeostasis and lead to accumulation of unfolded or misfolded proteins in the aging brain. Alzheimer's and Parkinson's diseases or Friedreich ataxia are neurological diseases sharing, as a common denominator, production of abnormal proteins, mitochondrial dysfunction and oxidative stress, which contribute to the pathogenesis of these so called "protein conformational diseases". The central nervous system has evolved the conserved mechanism of unfolded protein response to cope with the accumulation of misfolded proteins. As one of the main intracellular redox systems involved in neuroprotection, the vitagene system is emerging as a neurohormetic potential target for novel cytoprotective interventions. Vitagenes encode for cytoprotective heat shock proteins (Hsp) Hsp70 and heme oxygenase-1, as well as thioredoxin reductase and sirtuins. Nutritional studies show that ageing in animals can be significantly influenced by dietary restriction. Thus, the impact of dietary factors on health and longevity is an increasingly appreciated area of research. Reducing energy intake by controlled caloric restriction or intermittent fasting increases lifespan and protects various tissues against disease. Genetics has revealed that ageing may be controlled by changes in intracellular NAD/NADH ratio regulating sirtuin, a group of proteins linked to aging, metabolism and stress tolerance in several organisms. Recent findings suggest that several phytochemicals exhibit biphasic dose responses on cells with low doses activating signaling pathways that result in increased expression of vitagenes encoding survival proteins, as in the case of the Keap1/Nrf2/ARE pathway activated by curcumin and NAD/NADH-sirtuin-1 activated by resveratrol. Consistently, the neuroprotective roles of dietary antioxidants including

  15. Optimized Affinity Capture of Yeast Protein Complexes.

    Science.gov (United States)

    LaCava, John; Fernandez-Martinez, Javier; Hakhverdyan, Zhanna; Rout, Michael P

    2016-01-01

    Here, we describe an affinity isolation protocol. It uses cryomilled yeast cell powder for producing cell extracts and antibody-conjugated paramagnetic beads for affinity capture. Guidelines for determining the optimal extraction solvent composition are provided. Captured proteins are eluted in a denaturing solvent (sodium dodecyl sulfate polyacrylamide gel electrophoresis sample buffer) for gel-based proteomic analyses. Although the procedures can be modified to use other sources of cell extract and other forms of affinity media, to date we have consistently obtained the best results with the method presented. PMID:27371596

  16. Affinization of category O for quantum groups

    CERN Document Server

    Young, C A S

    2012-01-01

    Let g be a simple Lie algebra. We consider the category O-hat of those modules over the affine quantum group Uq(g-hat) whose Uq(g)-weights have finite multiplicity and lie in a finite union of cones generated by negative roots. We show that many properties of the category of the finite-dimensional representations naturally extend to the category O-hat. In particular, we develop the theory of q-characters and define the minimal affinizations of parabolic Verma modules. In types ABCFG we classify these minimal affinizations and conjecture a Weyl denominator type formula for their characters.

  17. Corner Transfer Matrices and Quantum Affine Algebras

    CERN Document Server

    Foda, O E; Foda, Omar; Miwa, Tetsuji

    1992-01-01

    Let H be the corner-transfer-matrix Hamiltonian for the six-vertex model in the anti-ferroelectric regime. It acts on the infinite tensor product W = V . V . V ....., where is the 2-dimensional irreducible representation of the quantum affine sl(2). We observe that H is the derivation of quantum affine sl(2), and conjecture that the eigenvectors of H form the level-1 vacuum representation of quantum affine sl(2). We report on checks in support of our conjecture.

  18. Mycobacterium tuberculosis Mce3E suppresses host innate immune responses by targeting ERK1/2 signaling.

    Science.gov (United States)

    Li, Jie; Chai, Qi-Yao; Zhang, Yong; Li, Bing-Xi; Wang, Jing; Qiu, Xiao-Bo; Liu, Cui Hua

    2015-04-15

    Crucial to the pathogenesis of the tuberculosis (TB)-causing pathogen Mycobacterium tuberculosis is its ability to subvert host immune defenses to promote its intracellular survival. The mammalian cell entry protein 3E (Mce3E), located in the region of difference 15 of the M. tuberculosis genome and absent in Mycobacterium bovis bacillus Calmette-Guérin, has an essential role in facilitating the internalization of mammalian cells by mycobacteria. However, relatively little is known about the role of Mce3E in modulation of host innate immune responses. In this study, we demonstrate that Mce3E inhibits the activation of the ERK1/2 signaling pathway, leading to the suppression of Tnf and Il6 expression, and the promotion of mycobacterial survival within macrophages. Mce3E interacts and colocalizes with ERK1/2 at the endoplasmic reticulum in a DEF motif (an ERK-docking motif)-dependent manner, relocates ERK1/2 from cytoplasm to the endoplasmic reticulum, and finally reduces the association of ERK1/2 with MEK1 and blocks the nuclear translocation of phospho-ERK1/2. A DEF motif mutant form of Mce3E (F294A) loses its ability to suppress Tnf and Il6 expression and to promote intracellular survival of mycobacteria. Inhibition of the ERK1/2 pathway in macrophages using U0126, a specific inhibitor of the ERK pathway, also leads to the suppressed Tnf and Il6 expression and the enhanced intracellular survival of mycobacteria. Taken together, these results suggest that M. tuberculosis Mce3E exploits the ERK1/2 signaling pathway to suppress host innate immune responses, providing a potential Mce3E-ERK1/2 interface-based drug target against M. tuberculosis. PMID:25780035

  19. Targeting the Innate Immune Response to Improve Cardiac Graft Recovery after Heart Transplantation: Implications for the Donation after Cardiac Death.

    Science.gov (United States)

    Toldo, Stefano; Quader, Mohammed; Salloum, Fadi N; Mezzaroma, Eleonora; Abbate, Antonio

    2016-01-01

    Heart transplantation (HTx) is the ultimate treatment for end-stage heart failure. The number of patients on waiting lists for heart transplants, however, is much higher than the number of available organs. The shortage of donor hearts is a serious concern since the population affected by heart failure is constantly increasing. Furthermore, the long-term success of HTx poses some challenges despite the improvement in the management of the short-term complications and in the methods to limit graft rejection. Myocardial injury occurs during transplantation. Injury initiated in the donor as result of brain or cardiac death is exacerbated by organ procurement and storage, and is ultimately amplified by reperfusion injury at the time of transplantation. The innate immune system is a mechanism of first-line defense against pathogens and cell injury. Innate immunity is activated during myocardial injury and produces deleterious effects on the heart structure and function. Here, we briefly discuss the role of the innate immunity in the initiation of myocardial injury, with particular focus on the Toll-like receptors and inflammasome, and how to potentially expand the donor population by targeting the innate immune response.

  20. Targeting the Innate Immune Response to Improve Cardiac Graft Recovery after Heart Transplantation: Implications for the Donation after Cardiac Death

    Directory of Open Access Journals (Sweden)

    Stefano Toldo

    2016-06-01

    Full Text Available Heart transplantation (HTx is the ultimate treatment for end-stage heart failure. The number of patients on waiting lists for heart transplants, however, is much higher than the number of available organs. The shortage of donor hearts is a serious concern since the population affected by heart failure is constantly increasing. Furthermore, the long-term success of HTx poses some challenges despite the improvement in the management of the short-term complications and in the methods to limit graft rejection. Myocardial injury occurs during transplantation. Injury initiated in the donor as result of brain or cardiac death is exacerbated by organ procurement and storage, and is ultimately amplified by reperfusion injury at the time of transplantation. The innate immune system is a mechanism of first-line defense against pathogens and cell injury. Innate immunity is activated during myocardial injury and produces deleterious effects on the heart structure and function. Here, we briefly discuss the role of the innate immunity in the initiation of myocardial injury, with particular focus on the Toll-like receptors and inflammasome, and how to potentially expand the donor population by targeting the innate immune response.

  1. Breast Tumor Targetable Fe3O4 Embedded Thermo-Responsive Nanoparticles for Radiofrequency Assisted Drug Delivery.

    Science.gov (United States)

    Rejinold, N Sanoj; Thomas, Reju George; Muthiah, Muthunarayanan; Lee, Hwa Jeongong; Jeong, Yong Yeon; Park, In-kyu; Jayakumar, R

    2016-01-01

    Non-invasive radiofrequency (RF) frequency may be utilized as an energy source to activate thermo-responsive nanoparticles for the controlled local delivery of drugs to cancer cells. Herein, we demonstrate that 180 ± 20 nm sized curcumin encapsulated chitosan-graft-poly(N-vinyl caprolactam) nanoparticles containing iron oxide nanoparticles (Fe3O4-CRC-TRC-NPs) were selectively internalized in cancer cells in vivo. Using an RF treatment at 80 watts for 2 min, Fe3O4-CRC-TRC-NPs, dissipated heat energy of 42 degrees C, which is the lower critical solution temperature (LCST) of the chitosan-graft-poly(N-vinyl caprolactam), causing controlled curcumin release and apoptosis to cultured 4T1 breast cancer cells. Further, the tumor localization studies on orthotopic breast cancer model revealed that Fe3O4-CRC-TRC-NPs selectively accumulated at the primary tumor as confirmed by in vivo live imaging followed by ex vivo tissue imaging and HPLC studies. These initial results strongly support the development of RF assisted drug delivery from nanoparticles for improved tumor targeting for breast cancer treatment. PMID:27301171

  2. PARP inhibitor ABT-888 affects response of MDA-MB-231 cells to doxorubicin treatment, targeting Snail expression.

    Science.gov (United States)

    Mariano, Germano; Ricciardi, Maria Rosaria; Trisciuoglio, Daniela; Zampieri, Michele; Ciccarone, Fabio; Guastafierro, Tiziana; Calabrese, Roberta; Valentini, Elisabetta; Tafuri, Agostino; Del Bufalo, Donatella; Caiafa, Paola; Reale, Anna

    2015-06-20

    To overcome cancer cells resistance to pharmacological therapy, the development of new therapeutic approaches becomes urgent. For this purpose, the use of poly(ADP-ribose) polymerase (PARP) inhibitors in combination with other cytotoxic agents could represent an efficacious strategy. Poly(ADP-ribosyl)ation (PARylation) is a post-translational modification that plays a well characterized role in the cellular decisions of life and death. Recent findings indicate that PARP-1 may control the expression of Snail, the master gene of epithelial-mesenchymal transition (EMT). Snail is highly represented in different resistant tumors, functioning as a factor regulating anti-apoptotic programmes. MDA-MB-231 is a Snail-expressing metastatic breast cancer cell line, which exhibits chemoresistance properties when treated with damaging agents. In this study, we show that the PARP inhibitor ABT-888 was capable to modulate the MDA-MB-231 cell response to doxorubicin, leading to an increase in the rate of apoptosis. Our further results indicate that PARP-1 controlled Snail expression at transcriptional level in cells exposed to doxorubicin. Given the increasing interest in the employment of PARP inhibitors as chemotherapeutic adjuvants, our in vitro results suggest that one of the mechanisms through which PARP inhibition can chemosensitize cancer cells in vivo, is targeting Snail expression thus promoting apoptosis. PMID:25938539

  3. Expression and affinity purification of recombinant proteins from plants

    Science.gov (United States)

    Desai, Urvee A.; Sur, Gargi; Daunert, Sylvia; Babbitt, Ruth; Li, Qingshun

    2002-01-01

    With recent advances in plant biotechnology, transgenic plants have been targeted as an inexpensive means for the mass production of proteins for biopharmaceutical and industrial uses. However, the current plant purification techniques lack a generally applicable, economic, large-scale strategy. In this study, we demonstrate the purification of a model protein, beta-glucuronidase (GUS), by employing the protein calmodulin (CaM) as an affinity tag. In the proposed system, CaM is fused to GUS. In the presence of calcium, the calmodulin fusion protein binds specifically to a phenothiazine-modified surface of an affinity column. When calcium is removed with a complexing agent, e.g., EDTA, calmodulin undergoes a conformational change allowing the dissociation of the calmodulin-phenothiazine complex and, therefore, permitting the elution of the GUS-CaM fusion protein. The advantages of this approach are the fast, efficient, and economical isolation of the target protein under mild elution conditions, thus preserving the activity of the target protein. Two types of transformation methods were used in this study, namely, the Agrobacterium-mediated system and the viral-vector-mediated transformation system. Copyright 2002 Elsevier Science (USA).

  4. Affitins for protein purification by affinity magnetic fishing.

    Science.gov (United States)

    Fernandes, Cláudia S M; Dos Santos, Raquel; Ottengy, Stella; Viecinski, Aline Canani; Béhar, Ghislaine; Mouratou, Barbara; Pecorari, Frédéric; Roque, A Cecília A

    2016-07-29

    Currently most economical and technological bottlenecks in protein production are placed in the downstream processes. With the aim of increasing the efficiency and reducing the associated costs, various affinity ligands have been developed. Affitins are small, yet robust and easy to produce, proteins derived from the archaeal extremophilic "7kDa DNA-binding" protein family. By means of combinatorial protein engineering and ribosome display selection techniques, Affitins have shown to bind a diversity of targets. In this work, two previously developed Affitins (anti-lysozyme and anti-IgG) were immobilized onto magnetic particles to assess their potential for protein purification by magnetic fishing. The optimal lysozyme and human IgG binding conditions yielded 58mg lysozyme/g support and 165mgIgG/g support, respectively. The recovery of proteins was possible in high yield (≥95%) and with high purity, namely ≥95% and 81%, when recovering lysozyme from Escherichia coli supernatant and IgG from human plasma, respectively. Static binding studies indicated affinity constants of 5.0×10(4)M(-1) and 9.3×10(5)M(-1) for the anti-lysozyme and anti-IgG magnetic supports. This work demonstrated that Affitins, which can be virtually evolved for any protein of interest, can be coupled onto magnetic particles creating novel affinity adsorbents for purification by magnetic fishing. PMID:27342136

  5. Automorphisms in Birational and Affine Geometry

    CERN Document Server

    Ciliberto, Ciro; Flenner, Hubert; McKernan, James; Prokhorov, Yuri; Zaidenberg, Mikhail

    2014-01-01

    The main focus of this volume is on the problem of describing the automorphism groups of affine and projective varieties, a classical subject in algebraic geometry where, in both cases, the automorphism group is often infinite dimensional. The collection covers a wide range of topics and is intended for researchers in the fields of classical algebraic geometry and birational geometry (Cremona groups) as well as affine geometry with an emphasis on algebraic group actions and automorphism groups. It presents original research and surveys and provides a valuable overview of the current state of the art in these topics. Bringing together specialists from projective, birational algebraic geometry and affine and complex algebraic geometry, including Mori theory and algebraic group actions, this book is the result of ensuing talks and discussions from the conference “Groups of Automorphisms in Birational and Affine Geometry” held in October 2012, at the CIRM, Levico Terme, Italy. The talks at the conference high...

  6. Synthesis of a New Series of Bone Affinity Compounds

    Institute of Scientific and Technical Information of China (English)

    2006-01-01

    A new series of bone affinity compounds were synthesized by linking chrysophanol with 5-fluorouracil derivatives. Their bone affinity was established by hydroxyapafive (HA)affinity experiment in vitro, and their cytostatic effects were shown by the MTT assay.

  7. A MEMS Dielectric Affinity Glucose Biosensor

    OpenAIRE

    Xian HUANG; Li, SiQi; Davis, Erin; Li, Dachao; Wang, Qian; Lin, Qiao

    2013-01-01

    Continuous glucose monitoring (CGM) sensors based on affinity detection are desirable for long-term and stable glucose management. However, most affinity sensors contain mechanical moving structures and complex design in sensor actuation and signal readout, limiting their reliability in subcutaneously implantable glucose detection. We have previously demonstrated a proof-of-concept dielectric glucose sensor that measured pre-mixed glucose-sensitive polymer solutions at various glucose concent...

  8. On Affine Fusion and the Phase Model

    OpenAIRE

    Walton, Mark A.

    2012-01-01

    A brief review is given of the integrable realization of affine fusion discovered recently by Korff and Stroppel. They showed that the affine fusion of the $su(n)$ Wess-Zumino-Novikov-Witten (WZNW) conformal field theories appears in a simple integrable system known as the phase model. The Yang-Baxter equation leads to the construction of commuting operators as Schur polynomials, with noncommuting hopping operators as arguments. The algebraic Bethe ansatz diagonalizes them, revealing a connec...

  9. Purely affine elementary su(N) fusions

    OpenAIRE

    Rasmussen, Jorgen; Walton, Mark A.

    2001-01-01

    We consider three-point couplings in simple Lie algebras -- singlets in triple tensor products of their integrable highest weight representations. A coupling can be expressed as a linear combination of products of finitely many elementary couplings. This carries over to affine fusion, the fusion of Wess-Zumino-Witten conformal field theories, where the expressions are in terms of elementary fusions. In the case of su(4) it has been observed that there is a purely affine elementary fusion, i.e...

  10. Complete algebraic vector fields on affine surfaces

    OpenAIRE

    Kaliman, Shulim; Kutzschebauch, Frank; Leuenberger, Matthias

    2014-01-01

    Let $\\AAutH (X)$ be the subgroup of the group $\\AutH (X)$ of holomorphic automorphisms of a normal affine algebraic surface $X$ generated by elements of flows associated with complete algebraic vector fields. Our main result is a classification of all normal affine algebraic surfaces $X$ quasi-homogeneous under $\\AAutH (X)$ in terms of the dual graphs of the boundaries $\\bX \\setminus X$ of their SNC-completions $\\bX$.

  11. Fan affinity laws from a collision model

    International Nuclear Information System (INIS)

    The performance of a fan is usually estimated using hydrodynamical considerations. The calculations are long and involved and the results are expressed in terms of three affinity laws. In this paper we use kinetic theory to attack this problem. A hard sphere collision model is used, and subsequently a correction to account for the flow behaviour of air is incorporated. Our calculations prove the affinity laws and provide numerical estimates of the air delivery, thrust and drag on a rotating fan. (paper)

  12. Architecture of high-affinity unnatural-base DNA aptamers toward pharmaceutical applications

    OpenAIRE

    Ken-ichiro Matsunaga; Michiko Kimoto; Charlotte Hanson; Michael Sanford; Young, Howard A.; Ichiro Hirao

    2015-01-01

    We present a remodeling method for high-affinity unnatural-base DNA aptamers to augment their thermal stability and nuclease resistance, for use as drug candidates targeting specific proteins. Introducing a unique mini-hairpin DNA provides robust stability to unnatural-base DNA aptamers generated by SELEX using genetic alphabet expansion, without reducing their high affinity. By this method, >80% of the remodeled DNA aptamer targeting interferon-γ (K D of 33 pM) survived in human serum at 37 ...

  13. Advances in identification and validation of protein targets of natural products without chemical modification.

    Science.gov (United States)

    Chang, J; Kim, Y; Kwon, H J

    2016-05-01

    Covering: up to February 2016Identification of the target proteins of natural products is pivotal to understanding the mechanisms of action to develop natural products for use as molecular probes and potential therapeutic drugs. Affinity chromatography of immobilized natural products has been conventionally used to identify target proteins, and has yielded good results. However, this method has limitations, in that labeling or tagging for immobilization and affinity purification often result in reduced or altered activity of the natural product. New strategies have recently been developed and applied to identify the target proteins of natural products and synthetic small molecules without chemical modification of the natural product. These direct and indirect methods for target identification of label-free natural products include drug affinity responsive target stability (DARTS), stability of proteins from rates of oxidation (SPROX), cellular thermal shift assay (CETSA), thermal proteome profiling (TPP), and bioinformatics-based analysis of connectivity. This review focuses on and reports case studies of the latest advances in target protein identification methods for label-free natural products. The integration of newly developed technologies will provide new insights and highlight the value of natural products for use as biological probes and new drug candidates. PMID:26964663

  14. The connection between metal ion affinity and ligand affinity in integrin I domains

    DEFF Research Database (Denmark)

    Vorup-Jensen, Thomas; Waldron, TT; Astrof, N;

    2007-01-01

    Integrins are cell-surface heterodimeric proteins that mediate cell-cell, cell-matrix, and cell-pathogen interactions. Half of the known integrin alpha subunits contain inserted domains (I domains) that coordinate ligand through a metal ion. Although the importance of conformational changes within...... isolated I domains in regulating ligand binding has been reported, the relationship between metal ion binding affinity and ligand binding affinity has not been elucidated. Metal and ligand binding by several I domain mutants that are stabilized in different conformations are investigated using isothermal...... titration calorimetry and surface plasmon resonance studies. This work suggests an inverse relationship between metal ion affinity and ligand binding affinity (i.e. constructs with a high affinity for ligand exhibit a low affinity for metal). This trend is discussed in the context of structural studies...

  15. Transfer RNA Derived Small RNAs Targeting Defense Responsive Genes Are Induced during Phytophthora capsici Infection in Black Pepper (Piper nigrum L.).

    Science.gov (United States)

    Asha, Srinivasan; Soniya, Eppurath V

    2016-01-01

    Small RNAs derived from transfer RNAs were recently assigned as potential gene regulatory candidates for various stress responses in eukaryotes. In this study, we report on the cloning and identification of tRNA derived small RNAs from black pepper plants in response to the infection of the quick wilt pathogen, Phytophthora capsici. 5'tRFs cloned from black pepper were validated as highly expressed during P. capsici infection. A high-throughput systematic analysis of the small RNAome (sRNAome) revealed the predominance of 5'tRFs in the infected leaf and root. The abundance of 5'tRFs in the sRNAome and the defense responsive genes as their potential targets indicated their regulatory role during stress response in black pepper. The 5'Ala(CGC) tRF mediated cleavage was experimentally mapped at the tRF binding sites on the mRNA targets of Non-expresser of pathogenesis related protein (NPR1), which was down-regulated during pathogen infection. Comparative sRNAome further demonstrated sequence conservation of 5'Ala tRFs across the angiosperm plant groups, and many important genes in the defense response were identified in silico as their potential targets. Our findings uncovered the diversity, differential expression and stress responsive functional role of tRNA-derived small RNAs during Phytophthora infection in black pepper. PMID:27313593

  16. Transfer RNA Derived Small RNAs Targeting Defense Responsive Genes Are Induced during Phytophthora capsici Infection in Black Pepper (Piper nigrum L.).

    Science.gov (United States)

    Asha, Srinivasan; Soniya, Eppurath V

    2016-01-01

    Small RNAs derived from transfer RNAs were recently assigned as potential gene regulatory candidates for various stress responses in eukaryotes. In this study, we report on the cloning and identification of tRNA derived small RNAs from black pepper plants in response to the infection of the quick wilt pathogen, Phytophthora capsici. 5'tRFs cloned from black pepper were validated as highly expressed during P. capsici infection. A high-throughput systematic analysis of the small RNAome (sRNAome) revealed the predominance of 5'tRFs in the infected leaf and root. The abundance of 5'tRFs in the sRNAome and the defense responsive genes as their potential targets indicated their regulatory role during stress response in black pepper. The 5'Ala(CGC) tRF mediated cleavage was experimentally mapped at the tRF binding sites on the mRNA targets of Non-expresser of pathogenesis related protein (NPR1), which was down-regulated during pathogen infection. Comparative sRNAome further demonstrated sequence conservation of 5'Ala tRFs across the angiosperm plant groups, and many important genes in the defense response were identified in silico as their potential targets. Our findings uncovered the diversity, differential expression and stress responsive functional role of tRNA-derived small RNAs during Phytophthora infection in black pepper.

  17. Affine modifications and affine hypersurfaces with a very transitive automorphism group

    OpenAIRE

    Kaliman, Shulim; ZAIDENBERG, MIKHAIL

    1998-01-01

    We study a kind of modification of an affine domain which produces another affine domain. First appeared in passing in the basic paper of O. Zariski (1942), it was further considered by E.D. Davis (1967). The first named author applied its geometric counterpart to construct contractible smooth affine varieties non-isomorphic to Euclidean spaces. Here we provide certain conditions which guarantee preservation of the topology under a modification. As an application, we show that the group of bi...

  18. The landscape of host transcriptional response programs commonly perturbed by bacterial pathogens: towards host-oriented broad-spectrum drug targets.

    Directory of Open Access Journals (Sweden)

    Yared H Kidane

    Full Text Available BACKGROUND: The emergence of drug-resistant pathogen strains and new infectious agents pose major challenges to public health. A promising approach to combat these problems is to target the host's genes or proteins, especially to discover targets that are effective against multiple pathogens, i.e., host-oriented broad-spectrum (HOBS drug targets. An important first step in the discovery of such drug targets is the identification of host responses that are commonly perturbed by multiple pathogens. RESULTS: In this paper, we present a methodology to identify common host responses elicited by multiple pathogens. First, we identified host responses perturbed by each pathogen using a gene set enrichment analysis of publicly available genome-wide transcriptional datasets. Then, we used biclustering to identify groups of host pathways and biological processes that were perturbed only by a subset of the analyzed pathogens. Finally, we tested the enrichment of each bicluster in human genes that are known drug targets, on the basis of which we elicited putative HOBS targets for specific groups of bacterial pathogens. We identified 84 up-regulated and three down-regulated statistically significant biclusters. Each bicluster contained a group of pathogens that commonly dysregulated a group of biological processes. We validated our approach by checking whether these biclusters correspond to known hallmarks of bacterial infection. Indeed, these biclusters contained biological process such as inflammation, activation of dendritic cells, pro- and anti- apoptotic responses and other innate immune responses. Next, we identified biclusters containing pathogens that infected the same tissue. After a literature-based analysis of the drug targets contained in these biclusters, we suggested new uses of the drugs Anakinra, Etanercept, and Infliximab for gastrointestinal pathogens Yersinia enterocolitica, Helicobacter pylori kx2 strain, and enterohemorrhagic Escherichia

  19. OptMAVEn--a new framework for the de novo design of antibody variable region models targeting specific antigen epitopes.

    Directory of Open Access Journals (Sweden)

    Tong Li

    Full Text Available Antibody-based therapeutics provides novel and efficacious treatments for a number of diseases. Traditional experimental approaches for designing therapeutic antibodies rely on raising antibodies against a target antigen in an immunized animal or directed evolution of antibodies with low affinity for the desired antigen. However, these methods remain time consuming, cannot target a specific epitope and do not lead to broad design principles informing other studies. Computational design methods can overcome some of these limitations by using biophysics models to rationally select antibody parts that maximize affinity for a target antigen epitope. This has been addressed to some extend by OptCDR for the design of complementary determining regions. Here, we extend this earlier contribution by addressing the de novo design of a model of the entire antibody variable region against a given antigen epitope while safeguarding for immunogenicity (Optimal Method for Antibody Variable region Engineering, OptMAVEn. OptMAVEn simulates in silico the in vivo steps of antibody generation and evolution, and is capable of capturing the critical structural features responsible for affinity maturation of antibodies. In addition, a humanization procedure was developed and incorporated into OptMAVEn to minimize the potential immunogenicity of the designed antibody models. As case studies, OptMAVEn was applied to design models of neutralizing antibodies targeting influenza hemagglutinin and HIV gp120. For both HA and gp120, novel computational antibody models with numerous interactions with their target epitopes were generated. The observed rates of mutations and types of amino acid changes during in silico affinity maturation are consistent with what has been observed during in vivo affinity maturation. The results demonstrate that OptMAVEn can efficiently generate diverse computational antibody models with both optimized binding affinity to antigens and reduced

  20. Early secretory antigenic target protein-6/culture filtrate protein-10 fusion protein-specific Th1 and Th2 response and its diagnostic value in tuberculous pleural effusion

    Institute of Scientific and Technical Information of China (English)

    戈启萍

    2013-01-01

    Objective To detect the Th1 and Th2 cell percentage in pleural effusion mononuclear cells (PEMCs) stimulated by early secretory antigenic target protein-6 (ESAT-6) /culture filtrate protein-10 (CFP-10) fusion protein (E/C) with flow cytometry (FCM) ,and to explore the local antigen specific Th1 and Th2 response and

  1. Glycan-modified liposomes boost CD4+ and CD8+ T-cell responses by targeting DC-SIGN on dendritic cells

    NARCIS (Netherlands)

    W.W.J. Unger; A.J. van Beelen; S.C. Bruijns; M. Joshi; C.M. Fehres; L. van Bloois; M.I. Verstege; M. Ambrosini; H. Kalay; K. Nazmi; J.G. Bolscher; E. Hooiberg; T.D. de Gruijl; G. Storm; Y. van Kooyk

    2012-01-01

    Cancer immunotherapy requires potent tumor-specific CD8+ and CD4+ T-cell responses, initiated by dendritic cells (DCs). Tumor antigens can be specifically targeted to DCs in vivo by exploiting their expression of C-type lectin receptors (CLR), which bind carbohydrate structures on antigens, resultin

  2. ACREAGE RESPONSE TO THE TARGET PRICE AND SET-ASIDE PROVISIONS OF THE FOOD AND AGRICULTURE ACT OF 1977

    OpenAIRE

    Evans, Sam

    1980-01-01

    The Food and Agriculture Act of 1977 increased the influence of target prices on acreage allocation decisions Differences between target and market prices were highly correlated with rates of participation in recent grain set-aside programs But, target prices also encourage set-aside participants to increase acreage of the set-aside crop The net effect of a set-aside on acreage of a specific crop may, thus, be positive or negative. Deficiencies in the target price formula magnify the potentia...

  3. The Protein-Protein Interface Evolution Acts in a Similar Way to Antibody Affinity Maturation*

    OpenAIRE

    Li, Bohua; Zhao, Lei; Wang, Chong; Guo, Huaizu; Wu, Lan; Zhang, Xunming; Qian, Weizhu; Wang, Hao; Guo, Yajun

    2009-01-01

    Understanding the evolutionary mechanism that acts at the interfaces of protein-protein complexes is a fundamental issue with high interest for delineating the macromolecular complexes and networks responsible for regulation and complexity in biological systems. To investigate whether the evolution of protein-protein interface acts in a similar way as antibody affinity maturation, we incorporated evolutionary information derived from antibody affinity maturation with common simulation techniq...

  4. Endoplasmic Reticulum Stress and Bipolar Disorder - Almost Forgotten Therapeutic Drug Targets in the Unfolded Protein Response Pathway Revisited.

    Science.gov (United States)

    Bengesser, Susanne A; Fuchs, Robert; Lackner, Nina; Birner, Armin; Reininghaus, Bernd; Meier-Allard, Nathalie; Stracke, Anika; Kapfhammer, Hans-Peter; Reininghaus, Eva Z; Wallner-Liebmann, Sandra

    2016-01-01

    Bipolar Disorder (BD) is characterized by recurring mood swings, which are not completely understood yet. So far, it is an accepted theory that multiple factors contribute to pathogenesis of BD according to the vulnerability-stressmodel. This model combines on the one hand biological predisposing vulnerability, and on the other hand several chronic and acute stressful negative events as underlying mechanisms of BD. Recently, ER (Endoplasmic Reticulum) stress reached the spotlight of BD research again. The expression of the chaperone BiP (syn. GRP78/glucose-regulated protein, 78kDa), which is highly expressed in the Endoplasmic Reticulum (ER), is upregulated by different kinds of mood stabilizers. These results implied that the ER, an organelle which is prone towards different kinds of cellular stress, might be involved in the pathophysiology of BD. This hypothesis was further strengthened by hypothesis driven genetic association studies, which showed significant association of BiP promotor polymorphisms with BD. Also other ER-stress associated genes like XBP1 (X-box binding protein 1) or GRP94 (glucose-regulated protein, 94kDa, synonym for heat shock protein HSP90B1) were recently linked to BD in hypothesis driven gene association studies. In addition to the proteins mentioned before, our review focuses on further UPR (Unfolded Protein Response) related proteins associated with BD and raises the hypothesis that ER-stress may represent a common interface between BD and obesity which is overrepresented in BD patients. Finally, members of the UPR pathway are discussed as putative targets for mood stabilizers.

  5. Downregulation of chloroplast RPS1 negatively modulates nuclear heat-responsive expression of HsfA2 and its target genes in Arabidopsis.

    Directory of Open Access Journals (Sweden)

    Hai-Dong Yu

    Full Text Available Heat stress commonly leads to inhibition of photosynthesis in higher plants. The transcriptional induction of heat stress-responsive genes represents the first line of inducible defense against imbalances in cellular homeostasis. Although heat stress transcription factor HsfA2 and its downstream target genes are well studied, the regulatory mechanisms by which HsfA2 is activated in response to heat stress remain elusive. Here, we show that chloroplast ribosomal protein S1 (RPS1 is a heat-responsive protein and functions in protein biosynthesis in chloroplast. Knockdown of RPS1 expression in the rps1 mutant nearly eliminates the heat stress-activated expression of HsfA2 and its target genes, leading to a considerable loss of heat tolerance. We further confirm the relationship existed between the downregulation of RPS1 expression and the loss of heat tolerance by generating RNA interference-transgenic lines of RPS1. Consistent with the notion that the inhibited activation of HsfA2 in response to heat stress in the rps1 mutant causes heat-susceptibility, we further demonstrate that overexpression of HsfA2 with a viral promoter leads to constitutive expressions of its target genes in the rps1 mutant, which is sufficient to reestablish lost heat tolerance and recovers heat-susceptible thylakoid stability to wild-type levels. Our findings reveal a heat-responsive retrograde pathway in which chloroplast translation capacity is a critical factor in heat-responsive activation of HsfA2 and its target genes required for cellular homeostasis under heat stress. Thus, RPS1 is an essential yet previously unknown determinant involved in retrograde activation of heat stress responses in higher plants.

  6. ESAT-6 Targeting to DEC205+ Antigen Presenting Cells Induces Specific-T Cell Responses against ESAT-6 and Reduces Pulmonary Infection with Virulent Mycobacterium tuberculosis.

    Directory of Open Access Journals (Sweden)

    Aarón Silva-Sánchez

    Full Text Available Airways infection with Mycobacterium tuberculosis (Mtb is contained mostly by T cell responses, however, Mtb has developed evasion mechanisms which affect antigen presenting cell (APC maturation/recruitment delaying the onset of Ag-specific T cell responses. Hypothetically, bypassing the natural infection routes by delivering antigens directly to APCs may overcome the pathogen's naturally evolved evasion mechanisms, thus facilitating the induction of protective immune responses. We generated a murine monoclonal fusion antibody (α-DEC-ESAT to deliver Early Secretory Antigen Target (ESAT-6 directly to DEC205+ APCs and to assess its in vivo effects on protection associated responses (IFN-γ production, in vivo CTL killing, and pulmonary mycobacterial load. Treatment with α-DEC-ESAT alone induced ESAT-6-specific IFN-γ producing CD4+ T cells and prime-boost immunization prior to Mtb infection resulted in early influx (d14 post-infection and increased IFN-γ+ production by specific T cells in the lungs, compared to scarce IFN-γ production in control mice. In vivo CTL killing was quantified in relevant tissues upon transferring target cells loaded with mycobacterial antigens. During infection, α-DEC-ESAT-treated mice showed increased target cell killing in the lungs, where histology revealed cellular infiltrate and considerably reduced bacterial burden. Targeting the mycobacterial antigen ESAT-6 to DEC205+ APCs before infection expands specific T cell clones responsible for early T cell responses (IFN-γ production and CTL activity and substantially reduces lung bacterial burden. Delivering mycobacterial antigens directly to APCs provides a unique approach to study in vivo the role of APCs and specific T cell responses to assess their potential anti-mycobacterial functions.

  7. Role of T cell receptor affinity in the efficacy and specificity of adoptive T cell therapies

    Directory of Open Access Journals (Sweden)

    Jennifer D. Stone

    2013-08-01

    Full Text Available Over the last several years, there has been considerable progress in the treatment of cancer using gene modified adoptive T cell therapies. Two approaches have been used, one involving the introduction of a conventional alpha-beta T cell receptor (TCR against a pepMHC cancer antigen, and the second involving introduction of a chimeric antigen receptor (CAR consisting of a single-chain antibody as an Fv fragment (scFv linked to transmembrane and signaling domains. In this review, we focus on one aspect of TCR-mediated adoptive T cell therapies, the impact of the affinity of the alpha-beta TCR for the pepMHC cancer antigen on both efficacy and specificity. We discuss the advantages of higher affinity TCRs in mediating potent activity of CD4 T cells. This is balanced with the potential disadvantage of higher affinity TCRs in mediating greater self-reactivity against a wider range of structurally similar antigenic peptides, especially in synergy with the CD8 co-receptor. Both TCR affinity and target selection will influence potential safety issues. We suggest pre-clinical strategies that might be used to examine each TCR for possible on-target and off-target side effects due to self-reactivities, and to adjust TCR affinities accordingly.

  8. Cassia obtusifolia MetE as a cytosolic target for potassium isolespedezate, a leaf-opening factor of Cassia plants: target exploration by a compact molecular-probe strategy.

    Science.gov (United States)

    Ueda, Minoru; Manabe, Yoshiyuki; Otsuka, Yuki; Kanzawa, Nobuyuki

    2011-12-01

    Affinity chromatography by using ligand-immobilized bead technology is generally the first choice for target exploration of a bioactive ligand. However, when a ligand has comparatively low affinity against its target, serious difficulties will be raised in affinity-based target detection. We report here that the use of compact molecular probes (CMP) will be advantageous in such cases; it enables the retention of moderate affinity between the ligand and its target in contrast to immobilizing the ligand on affinity beads that will cause a serious drop in affinity to preclude target detection. In the CMP strategy, a CMP containing an azide handle is used for an initial affinity-based labeling of target, and subsequent tagging by CuAAC with a large FLAG tag will give a tagged target protein. By using the CMP strategy, we succeeded in the identification of Cassia obtusifolia MetE as a cytosolic target protein of potassium isolespedezate (1), a moderately bioactive ligand.

  9. T-cell memory responses elicited by yellow fever vaccine are targeted to overlapping epitopes containing multiple HLA-I and -II binding motifs.

    Directory of Open Access Journals (Sweden)

    Andréa Barbosa de Melo

    Full Text Available The yellow fever vaccines (YF-17D-204 and 17DD are considered to be among the safest vaccines and the presence of neutralizing antibodies is correlated with protection, although other immune effector mechanisms are known to be involved. T-cell responses are known to play an important role modulating antibody production and the killing of infected cells. However, little is known about the repertoire of T-cell responses elicited by the YF-17DD vaccine in humans. In this report, a library of 653 partially overlapping 15-mer peptides covering the envelope (Env and nonstructural (NS proteins 1 to 5 of the vaccine was utilized to perform a comprehensive analysis of the virus-specific CD4(+ and CD8(+ T-cell responses. The T-cell responses were screened ex-vivo by IFN-γ ELISPOT assays using blood samples from 220 YF-17DD vaccinees collected two months to four years after immunization. Each peptide was tested in 75 to 208 separate individuals of the cohort. The screening identified sixteen immunodominant antigens that elicited activation of circulating memory T-cells in 10% to 33% of the individuals. Biochemical in-vitro binding assays and immunogenetic and immunogenicity studies indicated that each of the sixteen immunogenic 15-mer peptides contained two or more partially overlapping epitopes that could bind with high affinity to molecules of different HLAs. The prevalence of the immunogenicity of a peptide in the cohort was correlated with the diversity of HLA-II alleles that they could bind. These findings suggest that overlapping of HLA binding motifs within a peptide enhances its T-cell immunogenicity and the prevalence of the response in the population. In summary, the results suggests that in addition to factors of the innate immunity, "promiscuous" T-cell antigens might contribute to the high efficacy of the yellow fever vaccines.

  10. "The face of ostracism": The impact of the social categorization on the thermal facial responses of the target and the observer.

    Science.gov (United States)

    Paolini, Daniele; Alparone, Francesca R; Cardone, Daniela; van Beest, Ilja; Merla, Arcangelo

    2016-01-01

    Ostracism has been shown to elicit pain in both the target and the observers. Two experiments investigated the autonomic thermal signature associated with an ostracism experience and assessed whether and how social categorization impacts the autonomic arousal of both the target and the observer. Autonomic response was assessed using thermal infrared imaging, recording facial temperature variation during an online game of ball toss (i.e., Cyberball). Social categorization was manipulated using a minimal group paradigm. The results show a more intense autonomic response during ostracism (vs. inclusion), marked by an increase in facial temperature in the nose and the perioral area. This autonomic response is stronger when individuals are ostracized by ingroup (vs. outgroup) members. Similar pattern of temperature variations emerge when individuals observe an ostracism episode involving ingroup members. Our findings advance the understanding of psycho-physiological mechanisms underlying the ostracism experience and emphasize the impact of social categorization in such mechanisms. PMID:26613387

  11. Affinity purification of aprotinin from bovine lung.

    Science.gov (United States)

    Xin, Yu; Liu, Lanhua; Chen, Beizhan; Zhang, Ling; Tong, Yanjun

    2015-05-01

    An affinity protocol for the purification of aprotinin from bovine lung was developed. To simulate the structure of sucrose octasulfate, a natural specific probe for aprotinin, the affinity ligand was composed of an acidic head and a hydrophobic stick, and was then linked with Sepharose. The sorbent was then subjected to adsorption analysis with pure aprotinin. The purification process consisted of one step of affinity chromatography and another step of ultrafiltration. Then purified aprotinin was subjected to sodium dodecyl sulfate polyacrylamide gel electrophoresis, trypsin inhibitor activity, gel-filtration, and thin-layer chromatography analysis. As calculated, the theoretical maximum adsorption (Qmax ) of the affinity sorbent was 25,476.0 ± 184.8 kallikrein inactivator unit/g wet gel; the dissociation constant of the complex "immobilized ligand-aprotinin" (Kd ) was 4.6 ± 0.1 kallikrein inactivator unit/mL. After the affinity separation of bovine lung aprotinin, reducing sodium dodecyl sulfate polyacrylamide gel electrophoresis analysis and gel-filtration chromatography revealed that the protein was a single polypeptide, and the purities were ∼ 97 and 100%, respectively; the purified peptide was also confirmed with aprotinin standard by gel-filtration chromatography and thin-layer chromatography. After the whole purification process, protein, and bioactivity recoveries were 2.2 and 92.6%, respectively; and the specific activity was up to 15,907.1 ± 10.2 kallikrein inactivator unit/mg. PMID:25677462

  12. Classification of neocortical interneurons using affinity propagation

    Science.gov (United States)

    Santana, Roberto; McGarry, Laura M.; Bielza, Concha; Larrañaga, Pedro; Yuste, Rafael

    2013-01-01

    In spite of over a century of research on cortical circuits, it is still unknown how many classes of cortical neurons exist. In fact, neuronal classification is a difficult problem because it is unclear how to designate a neuronal cell class and what are the best characteristics to define them. Recently, unsupervised classifications using cluster analysis based on morphological, physiological, or molecular characteristics, have provided quantitative and unbiased identification of distinct neuronal subtypes, when applied to selected datasets. However, better and more robust classification methods are needed for increasingly complex and larger datasets. Here, we explored the use of affinity propagation, a recently developed unsupervised classification algorithm imported from machine learning, which gives a representative example or exemplar for each cluster. As a case study, we applied affinity propagation to a test dataset of 337 interneurons belonging to four subtypes, previously identified based on morphological and physiological characteristics. We found that affinity propagation correctly classified most of the neurons in a blind, non-supervised manner. Affinity propagation outperformed Ward's method, a current standard clustering approach, in classifying the neurons into 4 subtypes. Affinity propagation could therefore be used in future studies to validly classify neurons, as a first step to help reverse engineer neural circuits. PMID:24348339

  13. Engineering an antibody with picomolar affinity to DOTA chelates of multiple radionuclides for pretargeted radioimmunotherapy and imaging

    Energy Technology Data Exchange (ETDEWEB)

    Orcutt, Kelly Davis; Slusarczyk, Adrian L. [Department of Chemical Engineering, Massachusetts Institute of Technology, Cambridge, MA 02139 (United States); Cieslewicz, Maryelise [Department of Biological Engineering, Massachusetts Institute of Technology, Cambridge, MA 02139 (United States); Ruiz-Yi, Benjamin [Department of Chemical Engineering, Massachusetts Institute of Technology, Cambridge, MA 02139 (United States); Bhushan, Kumar R. [Division of Hematology/Oncology, Beth Israel Deaconess Medical Center, Boston, MA 02215 (United States); Frangioni, John V. [Division of Hematology/Oncology, Beth Israel Deaconess Medical Center, Boston, MA 02215 (United States); Department of Radiology, Beth Israel Deaconess Medical Center, Boston, MA 02215 (United States); Wittrup, K. Dane, E-mail: wittrup@mit.ed [Department of Chemical Engineering, Massachusetts Institute of Technology, Cambridge, MA 02139 (United States); Department of Biological Engineering, Massachusetts Institute of Technology, Cambridge, MA 02139 (United States); Koch Institute for Integrative Cancer Research, Massachusetts Institute of Technology, Cambridge, MA 02139 (United States)

    2011-02-15

    Introduction: In pretargeted radioimmunotherapy (PRIT), a bifunctional antibody is administered and allowed to pre-localize to tumor cells. Subsequently, a chelated radionuclide is administered and captured by cell-bound antibody while unbound hapten clears rapidly from the body. We aim to engineer high-affinity binders to 1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid (DOTA) chelates for use in PRIT applications. Methods: We mathematically modeled antibody and hapten pharmacokinetics to analyze hapten tumor retention as a function of hapten binding affinity. Motivated by model predictions, we used directed evolution and yeast surface display to affinity mature the 2D12.5 antibody to DOTA, reformatted as a single chain variable fragment (scFv). Results: Modeling predicts that for high antigen density and saturating bsAb dose, a hapten-binding affinity of 100 pM is needed for near-maximal hapten retention. We affinity matured 2D12.5 with an initial binding constant of about 10 nM to DOTA-yttrium chelates. Affinity maturation resulted in a 1000-fold affinity improvement to biotinylated DOTA-yttrium, yielding an 8.2{+-}1.9 picomolar binder. The high-affinity scFv binds DOTA complexes of lutetium and gadolinium with similar picomolar affinity and indium chelates with low nanomolar affinity. When engineered into a bispecific antibody construct targeting carcinoembryonic antigen, pretargeted high-affinity scFv results in significantly higher tumor retention of a {sup 111}In-DOTA hapten compared to pretargeted wild-type scFv in a xenograft mouse model. Conclusions: We have engineered a versatile, high-affinity, DOTA-chelate-binding scFv. We anticipate it will prove useful in developing pretargeted imaging and therapy protocols to exploit the potential of a variety of radiometals.

  14. Engineering an antibody with picomolar affinity to DOTA chelates of multiple radionuclides for pretargeted radioimmunotherapy and imaging

    International Nuclear Information System (INIS)

    Introduction: In pretargeted radioimmunotherapy (PRIT), a bifunctional antibody is administered and allowed to pre-localize to tumor cells. Subsequently, a chelated radionuclide is administered and captured by cell-bound antibody while unbound hapten clears rapidly from the body. We aim to engineer high-affinity binders to 1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid (DOTA) chelates for use in PRIT applications. Methods: We mathematically modeled antibody and hapten pharmacokinetics to analyze hapten tumor retention as a function of hapten binding affinity. Motivated by model predictions, we used directed evolution and yeast surface display to affinity mature the 2D12.5 antibody to DOTA, reformatted as a single chain variable fragment (scFv). Results: Modeling predicts that for high antigen density and saturating bsAb dose, a hapten-binding affinity of 100 pM is needed for near-maximal hapten retention. We affinity matured 2D12.5 with an initial binding constant of about 10 nM to DOTA-yttrium chelates. Affinity maturation resulted in a 1000-fold affinity improvement to biotinylated DOTA-yttrium, yielding an 8.2±1.9 picomolar binder. The high-affinity scFv binds DOTA complexes of lutetium and gadolinium with similar picomolar affinity and indium chelates with low nanomolar affinity. When engineered into a bispecific antibody construct targeting carcinoembryonic antigen, pretargeted high-affinity scFv results in significantly higher tumor retention of a 111In-DOTA hapten compared to pretargeted wild-type scFv in a xenograft mouse model. Conclusions: We have engineered a versatile, high-affinity, DOTA-chelate-binding scFv. We anticipate it will prove useful in developing pretargeted imaging and therapy protocols to exploit the potential of a variety of radiometals.

  15. Signaling in the yeast pheromone response pathway: specific and high-affinity interaction of the mitogen-activated protein (MAP) kinases Kss1 and Fus3 with the upstream MAP kinase kinase Ste7.

    OpenAIRE

    Bardwell, L; Cook, J G; E. C. Chang; Cairns, B R; Thorner, J

    1996-01-01

    Kss1 and Fus3 are mitogen-activated protein kinases (MAPKs or ERKs), and Ste7 is their activating MAPK/ERK kinase (MEK), in the pheromone response pathway of Saccharomyces cerevisiae. To investigate the potential role of specific interactions between these enzymes during signaling, their ability to associate with each other was examined both in solution and in vivo. When synthesized by in vitro translation, Kss1 and Fus3 could each form a tight complex (Kd of approximately 5 nM) with Ste7 in ...

  16. Stepparents' Affinity-Seeking and Affinity-Maintaining Strategies with Stepchildren.

    Science.gov (United States)

    Ganong, Lawrence; Coleman, Marilyn; Fine, Mark; Martin, Patricia

    1999-01-01

    Examines the strategies that stepparents use to develop and maintain affinity with stepchildren and the effects that these strategies have on the development of stepparent-stepchildren relationships. Thirty-one affinity-seeking strategies are identified. Results show that dyadic activities worked best, but it is important that stepchildren…

  17. Differential affinity of mammalian histone H1 somatic subtypes for DNA and chromatin

    Directory of Open Access Journals (Sweden)

    Mora Xavier

    2007-05-01

    chromatin suggest that differential affinity could be functionally relevant and thus contribute to the functional differentiation of the subtypes. The conservation of the relative affinities for SAR and non-SAR DNA, in spite of a strong preference for SAR sequences, indicates that differential affinity alone cannot be responsible for the heterogeneous distribution of some subtypes in cell nuclei.

  18. Heparin-binding peptide as a novel affinity tag for purification of recombinant proteins.

    Science.gov (United States)

    Morris, Jacqueline; Jayanthi, Srinivas; Langston, Rebekah; Daily, Anna; Kight, Alicia; McNabb, David S; Henry, Ralph; Kumar, Thallapuranam Krishnaswamy Suresh

    2016-10-01

    Purification of recombinant proteins constitutes a significant part of the downstream processing in biopharmaceutical industries. Major costs involved in the production of bio-therapeutics mainly depend on the number of purification steps used during the downstream process. Affinity chromatography is a widely used method for the purification of recombinant proteins expressed in different expression host platforms. Recombinant protein purification is achieved by fusing appropriate affinity tags to either N- or C- terminus of the target recombinant proteins. Currently available protein/peptide affinity tags have proved quite useful in the purification of recombinant proteins. However, these affinity tags suffer from specific limitations in their use under different conditions of purification. In this study, we have designed a novel 34-amino acid heparin-binding affinity tag (HB-tag) for the purification of recombinant proteins expressed in Escherichia coli (E. coli) cells. HB-tag fused recombinant proteins were overexpressed in E. coli in high yields. A one-step heparin-Sepharose-based affinity chromatography protocol was developed to purify HB-fused recombinant proteins to homogeneity using a simple sodium chloride step gradient elution. The HB-tag has also been shown to facilitate the purification of target recombinant proteins from their 8 M urea denatured state(s). The HB-tag has been demonstrated to be successfully released from the fusion protein by an appropriate protease treatment to obtain the recombinant target protein(s) in high yields. Results of the two-dimensional NMR spectroscopy experiments indicate that the purified recombinant target protein(s) exist in the native conformation. Polyclonal antibodies raised against the HB-peptide sequence, exhibited high binding specificity and sensitivity to the HB-fused recombinant proteins (∼10 ng) in different crude cell extracts obtained from diverse expression hosts. In our opinion, the HB-tag provides a

  19. On Affine Fusion and the Phase Model

    Directory of Open Access Journals (Sweden)

    Mark A. Walton

    2012-11-01

    Full Text Available A brief review is given of the integrable realization of affine fusion discovered recently by Korff and Stroppel. They showed that the affine fusion of the su(n Wess-Zumino-Novikov-Witten (WZNW conformal field theories appears in a simple integrable system known as the phase model. The Yang-Baxter equation leads to the construction of commuting operators as Schur polynomials, with noncommuting hopping operators as arguments. The algebraic Bethe ansatz diagonalizes them, revealing a connection to the modular S matrix and fusion of the su(n WZNW model. The noncommutative Schur polynomials play roles similar to those of the primary field operators in the corresponding WZNW model. In particular, their 3-point functions are the su(n fusion multiplicities. We show here how the new phase model realization of affine fusion makes obvious the existence of threshold levels, and how it accommodates higher-genus fusion.

  20. Affinity chromatography of bacterial lactate dehydrogenases.

    Science.gov (United States)

    Kelly, N; Delaney, M; O'Carra, P

    1978-06-01

    The affinity system used was the immobilized oxamate derivative previously used to purify mammalian lactate dehydrogenases. The bacterial dehydrogenases specific for the L-stereoisomer of lactate behaved in the same way as the mammalian enzymes, binding strongly in the presence of NADH. The D-lactate-specific enzymes, however, did not show any biospecific affinity for this gel. The L-specific enzymes could be purified to homogeneity in one affinity-chromatographic step. The D-specific enzymes could be efficiently separated from the L-specific ones and could then be further purified on an immobilized NAD derivative. The mechanism of activation of the lactate dehydrogenase from Streptococcus faecalis by fructose 1,6-bisphosphate was investigated by using the immobilized oxamate gel. PMID:666726

  1. Parity-dependent recognition of DBL1X-3X suggests an important role of the VAR2CSA high-affinity CSA-binding region in the development of the humoral response against placental malaria.

    Science.gov (United States)

    Dechavanne, Sébastien; Srivastava, Anand; Gangnard, Stéphane; Nunes-Silva, Sofia; Dechavanne, Célia; Fievet, Nadine; Deloron, Philippe; Chêne, Arnaud; Gamain, Benoît

    2015-06-01

    Plasmodium falciparum multidomain protein VAR2CSA stands today as the leading vaccine candidate against pregnancy-associated malaria (PAM). Most of the studies aiming to decrypt how naturally acquired immunity develops have assessed the immune recognition of individual VAR2CSA Duffy-binding-like (DBL) domains, thus overlooking the presence of conformational epitopes resulting from the overall folding of the full-length protein. In order to characterize the development of humoral immunity toward VAR2CSA, we made use of a large cohort of 293 Senegalese pregnant women to assess the level of recognition by plasma IgG of the full-length VAR2CSA protein of the 3D7 parasite strain (3D7-VAR2CSA), the CSA-binding multidomains 3D7-DBL1X to -DBL3X (3D7-DBL1X-3X), and the CSA nonbinding multidomains 3D7-DBL4ε to -DBL6ε (3D7-DBL4ε-6ε), as well as individual 3D7-DBL domains. Our results revealed a parity-dependent recognition of the full-length 3D7-VAR2CSA and of the CSA-binding region, 3D7-DBL1X-3X. Indeed, multigravid women possess significantly higher levels of antibodies directed against these constructs than primigravidae. Our results suggest an important role of antibodies targeting the CSA-binding region in the development of immunity against PAM, therefore providing new insights on how natural protection might be acquired and further information for the design of VAR2CSA-based vaccines. PMID:25824842

  2. The Affine q-Schur algebra

    OpenAIRE

    Green, R. M.

    1997-01-01

    We introduce an analogue of the $q$-Schur algebra associated to Coxeter systems of type $\\hat A_{n-1}$. We give two constructions of this algebra. The first construction realizes the algebra as a certain endomorphism algebra arising from an affine Hecke algebra of type $\\hat A_{r-1}$, where $n \\geq r$. This generalizes the original $q$-Schur algebra as defined by Dipper and James, and the new algebra contains the ordinary $q$-Schur algebra and the affine Hecke algebra as subalgebras. Using th...

  3. Affine Projection Algorithm Using Regressive Estimated Error

    OpenAIRE

    Zhang, Shu; Zhi, Yongfeng

    2011-01-01

    An affine projection algorithm using regressive estimated error (APA-REE) is presented in this paper. By redefining the iterated error of the affine projection algorithm (APA), a new algorithm is obtained, and it improves the adaptive filtering convergence rate. We analyze the iterated error signal and the stability for the APA-REE algorithm. The steady-state weights of the APA-REE algorithm are proved to be unbiased and consist. The simulation results show that the proposed algorithm has a f...

  4. Control and estimation of piecewise affine systems

    CERN Document Server

    Xu, Jun

    2014-01-01

    As a powerful tool to study nonlinear systems and hybrid systems, piecewise affine (PWA) systems have been widely applied to mechanical systems. Control and Estimation of Piecewise Affine Systems presents several research findings relating to the control and estimation of PWA systems in one unified view. Chapters in this title discuss stability results of PWA systems, using piecewise quadratic Lyapunov functions and piecewise homogeneous polynomial Lyapunov functions. Explicit necessary and sufficient conditions for the controllability and reachability of a class of PWA systems are

  5. Affine Invariant Character Recognition by Progressive Removing

    Science.gov (United States)

    Iwamura, Masakazu; Horimatsu, Akira; Niwa, Ryo; Kise, Koichi; Uchida, Seiichi; Omachi, Shinichiro

    Recognizing characters in scene images suffering from perspective distortion is a challenge. Although there are some methods to overcome this difficulty, they are time-consuming. In this paper, we propose a set of affine invariant features and a new recognition scheme called “progressive removing” that can help reduce the processing time. Progressive removing gradually removes less feasible categories and skew angles by using multiple classifiers. We observed that progressive removing and the use of the affine invariant features reduced the processing time by about 60% in comparison to a trivial one without decreasing the recognition rate.

  6. Adsorption affinity of anions on metal oxyhydroxides

    Science.gov (United States)

    Pechenyuk, S. I.; Semushina, Yu. P.; Kuz'mich, L. F.

    2013-03-01

    The dependences of anion (phosphate, carbonate, sulfate, chromate, oxalate, tartrate, and citrate) adsorption affinity anions from geometric characteristics, acid-base properties, and complex forming ability are generalized. It is shown that adsorption depends on the nature of both the anions and the ionic medium and adsorbent. It is established that anions are generally grouped into the following series of adsorption affinity reduction: PO{4/3-}, CO{3/2-} > C2O{4/2-}, C(OH)(CH2)2(COO){3/3-}, (CHOH)2(COO){2/2-} > CrO{4/2-} ≫ SO{4/2-}.

  7. Periodic cyclic homology of affine Hecke algebras

    CERN Document Server

    Solleveld, Maarten

    2009-01-01

    This is the author's PhD-thesis, which was written in 2006. The version posted here is identical to the printed one. Instead of an abstract, the short list of contents: Preface 5 1 Introduction 9 2 K-theory and cyclic type homology theories 13 3 Affine Hecke algebras 61 4 Reductive p-adic groups 103 5 Parameter deformations in affine Hecke algebras 129 6 Examples and calculations 169 A Crossed products 223 Bibliography 227 Index 237 Samenvatting 245 Curriculum vitae 253

  8. Design of cyclic peptides that bind protein surfaces with antibody-like affinity.

    Science.gov (United States)

    Millward, Steven W; Fiacco, Stephen; Austin, Ryan J; Roberts, Richard W

    2007-09-21

    There is a pressing need for new molecular tools to target protein surfaces with high affinity and specificity. Here, we describe cyclic messenger RNA display with a trillion-member covalent peptide macrocycle library. Using this library, we have designed a number of high-affinity, redox-insensitive, cyclic peptides that target the signaling protein G alpha i1. In addition to cyclization, our library construction took advantage of an expanded genetic code, utilizing nonsense suppression to insert N-methylphenylalanine as a 21st amino acid. The designed macrocycles exhibit several intriguing features. First, the core motif seen in all of the selected variants is the same and shares an identical context with respect to the macrocyclic scaffold, consistent with the idea that selection simultaneously optimizes both the cyclization chemistry and the structural placement of the binding epitope. Second, detailed characterization of one molecule, cyclic G alpha i binding peptide (cycGiBP), demonstrates substantially enhanced proteolytic stability relative to that of the parent linear molecule. Third and perhaps most important, the cycGiBP peptide binds the target with very high affinity ( K i approximately 2.1 nM), similar to those of many of the best monoclonal antibodies and higher than that of the betagamma heterodimer, an endogenous G alpha i1 ligand. Overall the work provides a general route to design novel, low-molecular-weight, high-affinity ligands that target protein surfaces.

  9. Improving Binding Affinity and Selectivity of Computationally Designed Ligand-Binding Proteins Using Experiments.

    Science.gov (United States)

    Tinberg, Christine E; Khare, Sagar D

    2016-01-01

    The ability to de novo design proteins that can bind small molecules has wide implications for synthetic biology and medicine. Combining computational protein design with the high-throughput screening of mutagenic libraries of computationally designed proteins is emerging as a general approach for creating binding proteins with programmable binding modes, affinities, and selectivities. The computational step enables the creation of a binding site in a protein that otherwise does not (measurably) bind the intended ligand, and targeted mutagenic screening allows for validation and refinement of the computational model as well as provides orders-of-magnitude increases in the binding affinity. Deep sequencing of mutagenic libraries can provide insights into the mutagenic binding landscape and enable further affinity improvements. Moreover, in such a combined computational-experimental approach where the binding mode is preprogrammed and iteratively refined, selectivity can be achieved (and modulated) by the placement of specified amino acid side chain groups around the ligand in defined orientations. Here, we describe the experimental aspects of a combined computational-experimental approach for designing-using the software suite Rosetta-proteins that bind a small molecule of choice and engineering, using fluorescence-activated cell sorting and high-throughput yeast surface display, high affinity and ligand selectivity. We illustrated the utility of this approach by performing the design of a selective digoxigenin (DIG)-binding protein that, after affinity maturation, binds DIG with picomolar affinity and high selectivity over structurally related steroids. PMID:27094290

  10. Directed evolution of antibody fragments with monovalent femtomolar antigen-binding affinity.

    Science.gov (United States)

    Boder, E T; Midelfort, K S; Wittrup, K D

    2000-09-26

    Single-chain antibody mutants have been evolved in vitro with antigen-binding equilibrium dissociation constant K(d) = 48 fM and slower dissociation kinetics (half-time > 5 days) than those for the streptavidin-biotin complex. These mutants possess the highest monovalent ligand-binding affinity yet reported for an engineered protein by over two orders of magnitude. Optimal kinetic screening of randomly mutagenized libraries of 10(5)-10(7) yeast surface-displayed antibodies enabled a >1,000-fold decrease in the rate of dissociation after four cycles of affinity mutagenesis and screening. The consensus mutations are generally nonconservative by comparison with naturally occurring mouse Fv sequences and with residues that do not contact the fluorescein antigen in the wild-type complex. The existence of these mutants demonstrates that the antibody Fv architecture is not intrinsically responsible for an antigen-binding affinity ceiling during in vivo affinity maturation.

  11. NK1 receptor fused to beta-arrestin displays a single-component, high-affinity molecular phenotype

    DEFF Research Database (Denmark)

    Martini, Lene; Hastrup, Hanne; Holst, Birgitte;

    2002-01-01

    with low affinity against antagonists. In contrast, in the NK1-beta-arrestin1 fusion protein, all ligands bound with similar affinity independent of the choice of radioligand and with Hill coefficients near unity. We conclude that the NK1 receptor in complex with arrestin is in a high-affinity, stable......Arrestins are cytosolic proteins that, upon stimulation of seven transmembrane (7TM) receptors, terminate signaling by binding to the receptor, displacing the G protein and targeting the receptor to clathrin-coated pits. Fusion of beta-arrestin1 to the C-terminal end of the neurokinin NK1 receptor...... Gq/G11 and Gs pathways. The NK1-beta-arrestin1 fusion construct bound nonpeptide antagonists with increased affinity but surprisingly also bound two types of agonists, substance P and neurokinin A, with high, normal affinity. In the wild-type NK1 receptor, neurokinin A (NKA) competes for binding...

  12. Crossing Chris: Some Markerian Affinities

    Directory of Open Access Journals (Sweden)

    Adrian Martin

    2010-01-01

    -pagination:widow-orphan; font-size:11.0pt; font-family:"Calibri","sans-serif"; mso-ascii-font-family:Calibri; mso-ascii-theme-font:minor-latin; mso-fareast-font-family:"Times New Roman"; mso-fareast-theme-font:minor-fareast; mso-hansi-font-family:Calibri; mso-hansi-theme-font:minor-latin; mso-bidi-font-family:"Times New Roman"; mso-bidi-theme-font:minor-bidi;}

    Abstract (E: This essay creatively explores a group of artists, writers, and other special individuals whose work or life story can be described as having an intriguing affinity with the protean career of Chris Marker. Avoiding the ‘usual suspects’ (such as Godard or Sebald, it discusses gossip columnist Milt Machlin, record collector Harry Smith, painter Gianfranco Baruchello, writer-filmmaker Edgardo Cozarinsky, and several others. From this constellation, a particular view of Markerian poetics emerges, touching upon the meanings of anonymity, storytelling, history and archiving.

     

    Abstract (F: Cet essai brosse de manière créative le portrait d’un groupe d'artistes, d'écrivains et d'autres personnes particulières dont le travail ou la biographie peuvent être décrits comme montrant une étrange mais certaine connivence avec la carrière protéiforme de Chris Marker. Evitant les lieux communs (comme Godard ou Sebald, cet article trace des références moins attendues :

  13. Transcriptome and Degradome of microRNAs and Their Targets in Response to Drought Stress in the Plants of a Diploid and Its Autotetraploid Paulownia australis.

    Directory of Open Access Journals (Sweden)

    Suyan Niu

    Full Text Available MicroRNAs (miRNAs are small, non-coding RNAs that play vital roles in plant growth, development, and stress response. Increasing numbers of studies aimed at discovering miRNAs and analyzing their functions in plants are being reported. In this study, we investigated the effect of drought stress on the expression of miRNAs and their targets in plants of a diploid and derived autotetraploid Paulownia australis. Four small RNA (sRNA libraries and four degradome libraries were constructed from diploid and autotetraploid P. australis plants treated with either 75% or 25% relative soil water content. A total of 33 conserved and 104 novel miRNAs (processing precision value > 0.1 were identified, and 125 target genes were identified for 36 of the miRNAs by using the degradome sequencing. Among the identified miRNAs, 54 and 68 were differentially expressed in diploid and autotetraploid plants under drought stress (25% relative soil water content, respectively. The expressions of miRNAs and target genes were also validated by quantitative real-time PCR. The results showed that the relative expression trends of the randomly selected miRNAs were similar to the trends predicted by Illumina sequencing. And the correlations between miRNAs and their target genes were also analyzed. Furthermore, the functional analysis showed that most of these miRNAs and target genes were associated with plant development and environmental stress response. This study provided molecular evidence for the possible involvement of certain miRNAs in the drought response and/or tolerance in P. australis, and certain level of differential expression between diploid and autotetraploid plants.

  14. Transcriptome and Degradome of microRNAs and Their Targets in Response to Drought Stress in the Plants of a Diploid and Its Autotetraploid Paulownia australis.

    Science.gov (United States)

    Niu, Suyan; Wang, Yuanlong; Zhao, Zhenli; Deng, Minjie; Cao, Lin; Yang, Lu; Fan, Guoqiang

    2016-01-01

    MicroRNAs (miRNAs) are small, non-coding RNAs that play vital roles in plant growth, development, and stress response. Increasing numbers of studies aimed at discovering miRNAs and analyzing their functions in plants are being reported. In this study, we investigated the effect of drought stress on the expression of miRNAs and their targets in plants of a diploid and derived autotetraploid Paulownia australis. Four small RNA (sRNA) libraries and four degradome libraries were constructed from diploid and autotetraploid P. australis plants treated with either 75% or 25% relative soil water content. A total of 33 conserved and 104 novel miRNAs (processing precision value > 0.1) were identified, and 125 target genes were identified for 36 of the miRNAs by using the degradome sequencing. Among the identified miRNAs, 54 and 68 were differentially expressed in diploid and autotetraploid plants under drought stress (25% relative soil water content), respectively. The expressions of miRNAs and target genes were also validated by quantitative real-time PCR. The results showed that the relative expression trends of the randomly selected miRNAs were similar to the trends predicted by Illumina sequencing. And the correlations between miRNAs and their target genes were also analyzed. Furthermore, the functional analysis showed that most of these miRNAs and target genes were associated with plant development and environmental stress response. This study provided molecular evidence for the possible involvement of certain miRNAs in the drought response and/or tolerance in P. australis, and certain level of differential expression between diploid and autotetraploid plants. PMID:27388154

  15. Transcriptome and Degradome of microRNAs and Their Targets in Response to Drought Stress in the Plants of a Diploid and Its Autotetraploid Paulownia australis

    Science.gov (United States)

    Niu, Suyan; Wang, Yuanlong; Zhao, Zhenli; Deng, Minjie; Cao, Lin; Yang, Lu; Fan, Guoqiang

    2016-01-01

    MicroRNAs (miRNAs) are small, non-coding RNAs that play vital roles in plant growth, development, and stress response. Increasing numbers of studies aimed at discovering miRNAs and analyzing their functions in plants are being reported. In this study, we investigated the effect of drought stress on the expression of miRNAs and their targets in plants of a diploid and derived autotetraploid Paulownia australis. Four small RNA (sRNA) libraries and four degradome libraries were constructed from diploid and autotetraploid P. australis plants treated with either 75% or 25% relative soil water content. A total of 33 conserved and 104 novel miRNAs (processing precision value > 0.1) were identified, and 125 target genes were identified for 36 of the miRNAs by using the degradome sequencing. Among the identified miRNAs, 54 and 68 were differentially expressed in diploid and autotetraploid plants under drought stress (25% relative soil water content), respectively. The expressions of miRNAs and target genes were also validated by quantitative real-time PCR. The results showed that the relative expression trends of the randomly selected miRNAs were similar to the trends predicted by Illumina sequencing. And the correlations between miRNAs and their target genes were also analyzed. Furthermore, the functional analysis showed that most of these miRNAs and target genes were associated with plant development and environmental stress response. This study provided molecular evidence for the possible involvement of certain miRNAs in the drought response and/or tolerance in P. australis, and certain level of differential expression between diploid and autotetraploid plants. PMID:27388154

  16. Rapid purification of circular DNA by triplex-mediated affinity capture

    Energy Technology Data Exchange (ETDEWEB)

    Ji, Huamin (4817 Sheboygan Ave., Madison, WI 53705); Smith, Lloyd M. (1115 Amherst Dr., Madison, WI 53705)

    1997-01-01

    A single-step capture of a target supercoiled double-stranded DNA molecule is accomplished by forming a local triple-helix among two strands of the supercoiled circular DNA and an oligonucleotide probe. The oligonucleotide is bound to an immobilizing support which facilitates the immobilization and purification of target DNA molecules. Non-target DNA molecules and other contaminating cellular material are easily removed by washing. The triple-helical structure is destabilized by raising the pH, leaving purified target DNA in the supernatant and reusable affinity capture oligonucleotide secured to the immobilizing support.

  17. Rapid purification of circular DNA by triplex-mediated affinity capture

    Energy Technology Data Exchange (ETDEWEB)

    Ji, H.; Smith, L.M.

    1997-01-07

    A single-step capture of a target supercoiled double-stranded DNA molecule is accomplished by forming a local triple-helix among two strands of the supercoiled circular DNA and an oligonucleotide probe. The oligonucleotide is bound to an immobilizing support which facilitates the immobilization and purification of target DNA molecules. Non-target DNA molecules and other contaminating cellular material are easily removed by washing. The triple-helical structure is destabilized by raising the pH, leaving purified target DNA in the supernatant and reusable affinity capture oligonucleotide secured to the immobilizing support. 3 figs.

  18. Identification of pancreatic tumors in vivo with ligand-targeted, pH responsive mesoporous silica nanoparticles by multispectral optoacoustic tomography.

    Science.gov (United States)

    Gurka, Marie K; Pender, Dillon; Chuong, Phillip; Fouts, Benjamin L; Sobelov, Alexander; McNally, Molly W; Mezera, Megan; Woo, Shiao Y; McNally, Lacey R

    2016-06-10

    Despite significant efforts to translate nanotechnology for cancer application, lack of identification of biodistribution/accumulation of these nanovehicles in vivo remains a substantial barrier for successful implementation of theranostic nanoparticles in the clinic. The purpose of the study was to develop a tumor-targeted theranostic nanovehicle for pancreatic cancer detectable by multispectral optoacoustic tomography (MSOT). To improve the tumor specificity of our mesoporous silica nanoparticle (MSN), we utilized a dual targeting strategy: 1) an elevated tumor receptor, urokinase plasminogen activator receptor (UPAR), and 2) the acidic tumor microenvironment. The tumor specificity of the MSN particle was improved with the addition of both chitosan, targeting acidic pH, and urokinase plasminogen activator (UPA), targeting UPAR. Drug release assays confirmed pH responsive release of gemcitabine in vitro. The UPAR specific binding of MSN-UPA nanoparticles was confirmed by reduction in fluorescence signal following MSN-UPA nanoparticle treatment in UPAR positive cells blocked with a UPAR-blocking antibody. Based upon Indocyanine Green encapsulation within the nanoparticles, UPA ligand targeted MSNs demonstrated increased intensity compared to untargeted MSNs at both pH7.4 (7×) and 6.5 (20×); however the signal was much more pronounced at a pH of 6.5 using tissue phantoms (pmultispectral optoacoustic tomography (p<0.05) and confirmed by ex vivo analysis. By tracking in vivo nanoparticle biodistribution with MSOT, it was shown that pH responsive, ligand targeted MSNs preferentially bind to pancreatic tumors for payload delivery. PMID:26763377

  19. Targeted delivery and pH-responsive release of stereoisomeric anti-cancer drugs using β-cyclodextrin assemblied Fe3O4 nanoparticles

    Science.gov (United States)

    Wang, Congli; Huang, Lizhen; Song, Shengmei; Saif, Bassam; Zhou, Yehong; Dong, Chuan; Shuang, Shaomin

    2015-12-01

    The β-cyclodextrin assemblied magnetic Fe3O4 nanoparticles (β-CD-MNPs) were successfully fabricated via a layer-by-layer method. Possessing an average size 14 nm, good stability and super-paramagnetic response (Ms 64 emu/g), the resultant nanocomposites could be served as a versatile biocompatible platform for selective loading, targeted delivery and pH-responsive release of stereoisomeric doxorubicin (DOX) and epirubicin (EPI). 1H-nuclear magnetic resonance (1H NMR) and the computer simulation further give the evidence that partial anthracene ring of drug molecule is included by β-CD. In addition, non-toxic β-CD-MNPs have excellent biocompatibility on MCF-7 cells, and cellular uptake indicate that different amounts of DOX or EPI can be transported to targeting site and released from the internalized carriers. The results demonstrate that as-prepared β-CD-MNPs could be a very promising vehicle for DOX and EPI.

  20. Core-shell magnetite nanoparticles surface encapsulated with smart stimuli-responsive polymer: synthesis, characterization, and LCST of viable drug-targeting delivery system.

    Science.gov (United States)

    Zhang, J L; Srivastava, R S; Misra, R D K

    2007-05-22

    We describe here the synthesis of a novel magnetic drug-targeting carrier characterized by a core-shell structure. The core-shell carrier combines the advantages of a magnetic core and the stimuli-responsive property of the thermosensitive biodegradable polymer shell (e.g., an on-off mechanism responsive to external temperature change). The composite nanoparticles are approximately 8 nm in diameter with approximately 3 nm shell. The lower critical solution temperature (LCST) is approximately 38 degrees C as determined by UV-vis absorption spectroscopy. The carrier is composed of cross-linked dextran grafted with a poly(N-isopropylacrylamide-co-N,N-dimethylacrylamide) [dextran-g-poly(NIPAAm-co-DMAAm)] shell and superparamagnetic Fe3O4 core. Fourier transform infrared spectroscopy (FTIR) confirmed the composition of the carrier. The synthesized magnetic carrier system has potential applications in magnetic drug-targeting delivery and magnetic resonance imaging.

  1. Fan Affinity Laws from a Collision Model

    Science.gov (United States)

    Bhattacharjee, Shayak

    2012-01-01

    The performance of a fan is usually estimated using hydrodynamical considerations. The calculations are long and involved and the results are expressed in terms of three affinity laws. In this paper we use kinetic theory to attack this problem. A hard sphere collision model is used, and subsequently a correction to account for the flow behaviour…

  2. Classification of neocortical interneurons using affinity propagation

    Directory of Open Access Journals (Sweden)

    Roberto eSantana

    2013-12-01

    Full Text Available In spite of over a century of research on cortical circuits, it is still unknown how many classes of cortical neurons exist. Neuronal classification has been a difficult problem because it is unclear what a neuronal cell class actually is and what are the best characteristics are to define them. Recently, unsupervised classifications using cluster analysis based on morphological, physiological or molecular characteristics, when applied to selected datasets, have provided quantitative and unbiased identification of distinct neuronal subtypes. However, better and more robust classification methods are needed for increasingly complex and larger datasets. We explored the use of affinity propagation, a recently developed unsupervised classification algorithm imported from machine learning, which gives a representative example or exemplar for each cluster. As a case study, we applied affinity propagation to a test dataset of 337 interneurons belonging to four subtypes, previously identified based on morphological and physiological characteristics. We found that affinity propagation correctly classified most of the neurons in a blind, non-supervised manner. In fact, using a combined anatomical/physiological dataset, our algorithm differentiated parvalbumin from somatostatin interneurons in 49 out of 50 cases. Affinity propagation could therefore be used in future studies to validly classify neurons, as a first step to help reverse engineer neural circuits.

  3. Magneto-nanosensor platform for probing low-affinity protein-protein interactions and identification of a low-affinity PD-L1/PD-L2 interaction.

    Science.gov (United States)

    Lee, Jung-Rok; Bechstein, Daniel J B; Ooi, Chin Chun; Patel, Ashka; Gaster, Richard S; Ng, Elaine; Gonzalez, Lino C; Wang, Shan X

    2016-01-01

    Substantial efforts have been made to understand the interactions between immune checkpoint receptors and their ligands targeted in immunotherapies against cancer. To carefully characterize the complete network of interactions involved and the binding affinities between their extracellular domains, an improved kinetic assay is needed to overcome limitations with surface plasmon resonance (SPR). Here, we present a magneto-nanosensor platform integrated with a microfluidic chip that allows measurement of dissociation constants in the micromolar-range. High-density conjugation of magnetic nanoparticles with prey proteins allows multivalent receptor interactions with sensor-immobilized bait proteins, more closely mimicking natural-receptor clustering on cells. The platform has advantages over traditional SPR in terms of insensitivity of signal responses to pH and salinity, less consumption of proteins and better sensitivities. Using this platform, we characterized the binding affinities of the PD-1-PD-L1/PD-L2 co-inhibitory receptor system, and discovered an unexpected interaction between the two known PD-1 ligands, PD-L1 and PD-L2. PMID:27447090

  4. Therapeutic targeting of regulatory T cells enhances tumor-specific CD8+ T cell responses in Epstein–Barr virus associated nasopharyngeal carcinoma

    Energy Technology Data Exchange (ETDEWEB)

    Fogg, Mark [Department of Medicine, Brigham and Women' s Hospital (United States); Murphy, John R. [Departments of Medicine and Microbiology, Boston University School of Medicine, Boston, MA 02118 (United States); Lorch, Jochen; Posner, Marshall [Department of Adult Oncology, Dana-Farber Cancer Institute, Harvard Medical School, Boston, MA 02115 (United States); Wang, Fred, E-mail: fwang@research.bwh.harvard.edu [Department of Medicine, Brigham and Women' s Hospital (United States); Department of Microbiology and Immunobiology, Harvard Medical School, Boston, MA 02115 (United States)

    2013-07-05

    Epstein–Barr virus (EBV) is associated with multiple malignancies including nasopharyngeal carcinoma (NPC). In nasopharynx cancer, CD8+ T cells specific for EBV Nuclear Antigen-1 (EBNA-1) and Latent Membrane Protein 2 (LMP2) are important components of anti-tumor immunity since both are consistently expressed in NPC. We have previously shown that EBNA-1-specific CD8+ T cell responses were suppressed in NPC patients compared to healthy controls. We now find that CD8+ T cell responses specific for LMP2 are also abnormal in NPC patients, and both EBNA-1- and LMP2-specific responses are suppressed by regulatory T cells (Treg). EBNA-1 and LMP2-specific CD8+ T cell responses, as well as immune control of EBV-infected cells in vitro, could be restored by the depletion of Tregs and by use of a clinically approved drug targeting Tregs. Thus, in vivo modulation of Tregs may be an effective means of enhancing these anti-tumor immune responses in NPC patients. - Highlights: • Viral proteins are tumor antigens in Epstein–Barr virus associated Nasopharyngeal Carcinoma. • CD8+ T cell responses against EBV proteins EBNA-1 and LMP2 are suppressed in NPC patients. • T regulatory cells are responsible for suppressing EBV immunity in NPC patients. • Depletion of Tregs with Ontak can rescue EBV-specific CD8+ T cell responses in NPC patients. • This clinically approved drug may be effective for enhancing anti-tumor immunity in NPC patients.

  5. Genome-Wide Investigation of MicroRNAs and Their Targets in Response to Freezing Stress in Medicago sativa L., Based on High-Throughput Sequencing

    Directory of Open Access Journals (Sweden)

    Yongjun Shu

    2016-03-01

    Full Text Available Winter damage, especially in northern climates, is a major limitation of the utilization of perennial forages such as alfalfa. Therefore, improving freezing tolerance is imperative in alfalfa genetic breeding. However, freezing tolerance is a complex trait that is determined by many genes. To understand the complex regulation mechanisms of freezing tolerance in alfalfa, we performed small RNA sequencing analysis under cold (4° and freezing (−8° stress. The sequencing results revealed that 173 known, and 24 novel miRNAs were expressed, and that the expression of 35 miRNAs was affected by cold and/or freezing stress. Meanwhile, 105 target genes cleaved by these miRNAs were characterized by degradome sequencing. These targets were associated with biological regulation, cellular processes, metabolic processes, and response to stress. Interestingly, most of them were characterized as transcription factors (TFs, including auxin response factors, SBP, NAC, AP2/ERF, and GRF, which play important roles in plant abiotic responses. In addition, important miRNAs and mRNAs involved in nodulation were also identified, for example, the relationship between miR169 and the TF CCAAT (also named as NF-YA/HAP2, which suggested that nodulation has an important function in freezing tolerance in alfalfa. Our results provide valuable information to help determine the molecular mechanisms of freezing tolerance in alfalfa, which will aid the application of these miRNAs and their targets in the improvement of freezing tolerance in alfalfa and related plants.

  6. 18F-FDG PET/CT for Monitoring the Response of Breast Cancer to miR-143-Based Therapeutics by Targeting Tumor Glycolysis

    Science.gov (United States)

    Miao, Ying; Zhang, Ling-fei; Guo, Rui; Liang, Sheng; Zhang, Min; Shi, Shuo; Shang-Guan, Cheng-fang; Liu, Mo-fang; Li, Biao

    2016-01-01

    Increased glucose utilization is a hallmark of cancer, and tumor metabolism is emerging as anticancer target for therapeutic intervention. Triple-negative breast cancers TNBC are highly glycolytic and show poor clinical outcomes. We previously identified hexokinase 2, the major glycolytic enzyme, as a target gene of miR-143 in TNBC. Here, we developed a therapeutic formulation using cholesterol-modified miR-143 agomir encapsulated in a neutral lipid-based delivery agent that blocked tumor growth and glucose metabolism in TNBC tumor-bearing mice when administered systemically. The antioncogenic effects were accompanied by a reduction in the direct target hexokinase 2 and [18F]-fluorodeoxyglucose (18F-FDG) uptake based on positron emission tomography/computed tomography. Treatment with miR-143 formulation has minimal toxic effects and mice tolerated it well. Thus, we demonstrated that miR-143 is a robust inhibitor of the Warburg effect and an effective therapeutic target for TNBC. In addition, 18F-FDG positron emission tomography/computed tomography can be used to specifically monitor the response of TNBC to miR-143-based therapeutics by targeting tumor glycolysis. PMID:27574783

  7. Targeting the tumor-draining area : local immunotherapy and its effect on the systemic T cell response

    NARCIS (Netherlands)

    Herbert-Fransen, Marieke Fernande

    2012-01-01

    This dissertation deals with the role of local immune stimulation in the lymph node and tumor microenvironment and its effect on systemic CD8+ T cell responses, in particular the anti-tumor CD8+ T cell responses. In chapter 2 the use of a slow-release system is described to deliver the immune-acti

  8. Binding Affinity of Glycoconjugates to BACILLUS Spores and Toxins

    Science.gov (United States)

    Rasol, Aveen; Eassa, Souzan; Tarasenko, Olga

    2010-04-01

    Early recognition of Bacillus cereus group species is important since they can cause food-borne illnesses and deadly diseases in humans. Glycoconjugates (GCs) are carbohydrates covalently linked to non-sugar moieties including lipids, proteins or other entities. GCs are involved in recognition and signaling processes intrinsic to biochemical functions in cells. They also stimulate cell-cell adhesion and subsequent recognition and activation of receptors. We have demonstrated that GCs are involved in Bacillus cereus spore recognition. In the present study, we have investigated whether GCs possess the ability to bind and recognize B. cereus spores and Bacillus anthracis recombinant single toxins (sTX) and complex toxins (cTX). The affinity of GCs to spores + sTX and spores + cTX toxins was studied in the binding essay. Our results demonstrated that GC9 and GC10 were able to selectively bind to B. cereus spores and B. anthracis toxins. Different binding affinities for GCs were found toward Bacillus cereus spores + sTX and spores + cTX. Dilution of GCs does not impede the recognition and binding. Developed method provides a tool for simultaneous recognition and targeting of spores, bacteria toxins, and/or other entities.

  9. Detection-Guided Fast Affine Projection Channel Estimator for Speech Applications

    Directory of Open Access Journals (Sweden)

    Yan Wu Jennifer

    2007-04-01

    Full Text Available In various adaptive estimation applications, such as acoustic echo cancellation within teleconferencing systems, the input signal is a highly correlated speech. This, in general, leads to extremely slow convergence of the NLMS adaptive FIR estimator. As a result, for such applications, the affine projection algorithm (APA or the low-complexity version, the fast affine projection (FAP algorithm, is commonly employed instead of the NLMS algorithm. In such applications, the signal propagation channel may have a relatively low-dimensional impulse response structure, that is, the number m of active or significant taps within the (discrete-time modelled channel impulse response is much less than the overall tap length n of the channel impulse response. For such cases, we investigate the inclusion of an active-parameter detection-guided concept within the fast affine projection FIR channel estimator. Simulation results indicate that the proposed detection-guided fast affine projection channel estimator has improved convergence speed and has lead to better steady-state performance than the standard fast affine projection channel estimator, especially in the important case of highly correlated speech input signals.

  10. DNA Vaccines Encoding Antigen Targeted to MHC Class II Induce Influenza-Specific CD8(+) T Cell Responses, Enabling Faster Resolution of Influenza Disease.

    Science.gov (United States)

    Lambert, Laura; Kinnear, Ekaterina; McDonald, Jacqueline U; Grodeland, Gunnveig; Bogen, Bjarne; Stubsrud, Elisabeth; Lindeberg, Mona M; Fredriksen, Agnete Brunsvik; Tregoning, John S

    2016-01-01

    Current influenza vaccines are effective but imperfect, failing to cover against emerging strains of virus and requiring seasonal administration to protect against new strains. A key step to improving influenza vaccines is to improve our understanding of vaccine-induced protection. While it is clear that antibodies play a protective role, vaccine-induced CD8(+) T cells can improve protection. To further explore the role of CD8(+) T cells, we used a DNA vaccine that encodes antigen dimerized to an immune cell targeting module. Immunizing CB6F1 mice with the DNA vaccine in a heterologous prime-boost regime with the seasonal protein vaccine improved the resolution of influenza disease compared with protein alone. This improved disease resolution was dependent on CD8(+) T cells. However, DNA vaccine regimes that induced CD8(+) T cells alone were not protective and did not boost the protection provided by protein. The MHC-targeting module used was an anti-I-E(d) single chain antibody specific to the BALB/c strain of mice. To test the role of MHC targeting, we compared the response between BALB/c, C57BL/6 mice, and an F1 cross of the two strains (CB6F1). BALB/c mice were protected, C57BL/6 were not, and the F1 had an intermediate phenotype; showing that the targeting of antigen is important in the response. Based on these findings, and in agreement with other studies using different vaccines, we conclude that, in addition to antibody, inducing a protective CD8 response is important in future influenza vaccines. PMID:27602032

  11. Congophilicity (Congo red affinity) of different beta2-microglobulin conformations characterized by dye affinity capillary electrophoresis

    DEFF Research Database (Denmark)

    Heegaard, N H; Sen, J W; Nissen, Mogens Holst

    2000-01-01

    The amyloidogenic protein beta-microglobulin was characterized by affinity capillary electrophoresis (CE). CE could separate conformational variants of beta2-microglobulin and with the amyloid-specific dye Congo red as a buffer additive it was possible to measure different Congo red-affinities of......The amyloidogenic protein beta-microglobulin was characterized by affinity capillary electrophoresis (CE). CE could separate conformational variants of beta2-microglobulin and with the amyloid-specific dye Congo red as a buffer additive it was possible to measure different Congo red......-affinities of native and abnormally folded beta2-microglobulin. We find that native beta2-microglobulin has an intermediate affinity for Congo red at pH 7.3 and that binding involves electrostatic interactions. The conformational variant of beta2-microglobulin that appears in acetonitrile solutions binds Congo red...... more strongly. Affinity CE using Congo red as a buffer additive is a new, simple, fast, and quantitative micromethod for the characterization of soluble conformational intermediates of amyloidogenic proteins....

  12. Evaluation of Efficiency Activities in the Industrial Sector Undertaken in Response to Greenhouse Gas Emission Reduction Targets

    Energy Technology Data Exchange (ETDEWEB)

    Price, Lynn; de la Rue du Can, Stephane; Lu, Hongyou; Horvath, Arpad

    2010-05-21

    The 2006 California Global Warming Solutions Act calls for reducing greenhouse gas (GHG) emissions to 1990 levels by 2020. Meeting this target will require action from all sectors of the California economy, including industry. The industrial sector consumes 25% of the energy used and emits 28% of the carbon dioxide (CO{sub 2}) produced in the state. Many countries around the world have national-level GHG reduction or energy-efficiency targets, and comprehensive programs focused on implementation of energy efficiency and GHG emissions mitigation measures in the industrial sector are essential for achieving their goals. A combination of targets and industry-focused supporting programs has led to significant investments in energy efficiency as well as reductions in GHG emissions within the industrial sectors in these countries. This project has identified program and policies that have effectively targeted the industrial sector in other countries to achieve real energy and CO{sub 2} savings. Programs in Ireland, France, The Netherlands, Denmark, and the UK were chosen for detailed review. Based on the international experience documented in this report, it is recommended that companies in California's industrial sector be engaged in a program to provide them with support to meet the requirements of AB32, The Global Warming Solution Act. As shown in this review, structured programs that engage industry, require members to evaluate their potential efficiency measures, plan how to meet efficiency or emissions reduction goals, and provide support in achieving the goals, can be quite effective at assisting companies to achieve energy efficiency levels beyond those that can be expected to be achieved autonomously.

  13. Down-regulation of Akt/mammalian target of rapamycin signaling pathway in response to myostatin overexpression in skeletal muscle.

    OpenAIRE

    Amirouche, Adel; Durieux, Anne-Cécile; Banzet, Sébastien; Koulmann, Nathalie; Bonnefoy, Régis; Mouret, Catherine; Bigard, Xavier; Peinnequin, André; Freyssenet, Damien

    2009-01-01

    Myostatin, a member of the TGF-beta family, has been identified as a master regulator of embryonic myogenesis and early postnatal skeletal muscle growth. However, cumulative evidence also suggests that alterations in skeletal muscle mass are associated with dysregulation in myostatin expression and that myostatin may contribute to muscle mass loss in adulthood. Two major branches of the Akt pathway are relevant for the regulation of skeletal muscle mass, the Akt/mammalian target of rapamycin ...

  14. Identification of putative TAL effector targets of the citrus canker pathogens shows functional convergence underlying disease development and defense response

    OpenAIRE

    Pereira, Andre LA; Carazzolle, Marcelo F.; Abe, Valeria Y; de Oliveira, Maria LP; Domingues, Mariane N; Silva, Jaqueline C; Cernadas, Raul A; Benedetti, Celso E.

    2014-01-01

    Background Transcriptional activator-like (TAL) effectors, formerly known as the AvrBs3/PthA protein family, are DNA-binding effectors broadly found in Xanthomonas spp. that transactivate host genes upon injection via the bacterial type three-secretion system. Biologically relevant targets of TAL effectors, i.e. host genes whose induction is vital to establish a compatible interaction, have been reported for xanthomonads that colonize rice and pepper; however, citrus genes modulated by the TA...

  15. Metabolomics Reveals Metabolic Targets and Biphasic Responses in Breast Cancer Cells Treated by Curcumin Alone and in Association with Docetaxel

    OpenAIRE

    Bayet-Robert, Mathilde; Morvan, Daniel

    2013-01-01

    Background Curcumin (CUR) has deserved extensive research due to its anti-inflammatory properties, of interest in human diseases including cancer. However, pleiotropic even paradoxical responses of tumor cells have been reported, and the mechanisms of action of CUR remain uncompletely elucidated. Methodology/Principal Findings 1H-NMR spectroscopy-based metabolomics was applied to get novel insight into responses of MCF7 and MDA-MB-231 breast cancer cells to CUR alone, and MCF7 cells to CUR in...

  16. A chirality change in XPC- and Sfi1-derived peptides affects their affinity for centrin.

    Science.gov (United States)

    Grecu, Dora; Irudayaraj, Victor Paul Raj; Martinez-Sanz, Juan; Mallet, Jean-Maurice; Assairi, Liliane

    2016-04-01

    The Ca(2+)-binding protein centrin binds to a hydrophobic motif (W(1)xxL(4)xxxL(8)) included in the sequence of several cellular targets: XPC (xeroderma pigmentosum group C protein), Sfi1 (suppressor of fermentation-induced loss of stress resistance protein1), and Sac3 [the central component of the transcription and mRNA export (TREX-2) complex]. However, centrin binding occurs in a reversed orientation (L(8)xxxL(4)xxW(1)) for Sfi1 and Sac3 compared with XPC. Because D-peptides have been investigated for future therapeutic use, we analyzed their centrin-binding properties. Their affinity for centrin was measured using isothermal titration calorimetry. The chirality change in the target-derived peptides affected their ability to bind centrin in a specific manner depending on the sequence orientation of the centrin-binding motif. In contrast to L-XPC-P10, D-XPC-P10 bound C-HsCen1 in a Ca(2+)-dependent manner and to a lesser extent. D-XPC-P10 exhibited a reduced affinity for C-HsCen1 (Ka=0.064 × 10(6) M(-1)) by a factor of 2000 compared with L-XPC-P10 (Ka=132 × 10(6) M(-1)). D-peptides have a lower affinity than L-peptides for centrin, and the strength of this affinity depends on the sequence orientation of the target-derived peptides. The residual affinity observed for D-XPC suggests that the use of d-peptides represents a promising strategy for inhibiting centrin binding to its targets. PMID:26923803

  17. A chirality change in XPC- and Sfi1-derived peptides affects their affinity for centrin.

    Science.gov (United States)

    Grecu, Dora; Irudayaraj, Victor Paul Raj; Martinez-Sanz, Juan; Mallet, Jean-Maurice; Assairi, Liliane

    2016-04-01

    The Ca(2+)-binding protein centrin binds to a hydrophobic motif (W(1)xxL(4)xxxL(8)) included in the sequence of several cellular targets: XPC (xeroderma pigmentosum group C protein), Sfi1 (suppressor of fermentation-induced loss of stress resistance protein1), and Sac3 [the central component of the transcription and mRNA export (TREX-2) complex]. However, centrin binding occurs in a reversed orientation (L(8)xxxL(4)xxW(1)) for Sfi1 and Sac3 compared with XPC. Because D-peptides have been investigated for future therapeutic use, we analyzed their centrin-binding properties. Their affinity for centrin was measured using isothermal titration calorimetry. The chirality change in the target-derived peptides affected their ability to bind centrin in a specific manner depending on the sequence orientation of the centrin-binding motif. In contrast to L-XPC-P10, D-XPC-P10 bound C-HsCen1 in a Ca(2+)-dependent manner and to a lesser extent. D-XPC-P10 exhibited a reduced affinity for C-HsCen1 (Ka=0.064 × 10(6) M(-1)) by a factor of 2000 compared with L-XPC-P10 (Ka=132 × 10(6) M(-1)). D-peptides have a lower affinity than L-peptides for centrin, and the strength of this affinity depends on the sequence orientation of the target-derived peptides. The residual affinity observed for D-XPC suggests that the use of d-peptides represents a promising strategy for inhibiting centrin binding to its targets.

  18. Improved native affinity purification of RNA.

    Science.gov (United States)

    Batey, Robert T; Kieft, Jeffrey S

    2007-08-01

    RNA biochemical or structural studies often require an RNA sample that is chemically pure, and most protocols for its in vitro production use denaturing polyacrylamide gel electrophoresis to achieve this. Unfortunately, many RNAs do not quantitatively refold into an active conformation after denaturation, creating significant problems for downstream characterization or use. In addition, this traditional purification method is not amenable to studies demanding high-throughput RNA production. Recently, we presented the first general method for producing almost any RNA sequence that employs an affinity tag that is removed during the purification process. Because technical difficulties prevented application of this method to many RNAs, we have developed an improved version that utilizes a different activatable ribozyme and affinity tag that are considerably more robust, rapid, and broadly applicable. PMID:17548432

  19. AFFINITY OF LIGNIN PREPARATIONS TOWARDS GENOTOXIC COMPOUNDS

    Directory of Open Access Journals (Sweden)

    Božena Košíková

    2009-02-01

    Full Text Available The carcinogenicity and mutagenicity of chemicals may be modulated by other chemicals, including those prepared by organic synthesis. Consid-ering the several drawbacks of synthetic compounds vis-a-vis the human organism, the lignin biomass component was examined for this purpose. The binding affinity of lignin samples prepared by chemical and biological modification of lignin products derived from chemical wood treatment towards for N-nitrosodiethylamine (NDA was examined. The protective role of the lignin samples against carcinogenesis was tested on a well-known model carcinogen, N-methyl-N´-nitro-N-nitrosoguanidine (MNNG. The observed ability of a series of lignin preparations to reduce alkylation damage of deoxyribonucleic acid (DNA on hamster cells in vitro could be explained by their affinity to bind N-nitrosoamines. The results indicate that lignin has potential to protect living organisms against damaging effects of different genotoxicants.

  20. Local structure of self-affine sets

    CERN Document Server

    Bandt, Christoph

    2011-01-01

    The structure of a self-similar set with open set condition does not change under magnification. For self-affine sets the situation is completely different. We consider planar self-affine Cantor sets E of the type studied by Bedford, McMullen, Gatzouras and Lalley, for which the projection onto the horizontal axis is an interval. We show that within small square neighborhoods of almost each point x in E, with respect to many product measures on address space, E is well approximated by product sets of an interval and a Cantor set. Even though E is totally disconnected, the limit sets have the product structure with interval fibres, reminiscent to the view of attractors of chaotic differentiable dynamical systems.

  1. Recent Results Regarding Affine Quantum Gravity

    CERN Document Server

    Klauder, John R

    2012-01-01

    Recent progress in the quantization of nonrenormalizable scalar fields has found that a suitable non-classical modification of the ground state wave function leads to a result that eliminates term-by-term divergences that arise in a conventional perturbation analysis. After a brief review of both the scalar field story and the affine quantum gravity program, examination of the procedures used in the latter surprisingly shows an analogous formulation which already implies that affine quantum gravity is not plagued by divergences that arise in a standard perturbation study. Additionally, guided by the projection operator method to deal with quantum constraints, trial reproducing kernels are introduced that satisfy the diffeomorphism constraints. Furthermore, it is argued that the trial reproducing kernels for the diffeomorphism constraints may also satisfy the Hamiltonian constraint as well.

  2. Derivation of injury-responsive dendritic cells for acute brain targeting and therapeutic protein delivery in the stroke-injured rat.

    Directory of Open Access Journals (Sweden)

    Nathan C Manley

    Full Text Available Research with experimental stroke models has identified a wide range of therapeutic proteins that can prevent the brain damage caused by this form of acute neurological injury. Despite this, we do not yet have safe and effective ways to deliver therapeutic proteins to the injured brain, and this remains a major obstacle for clinical translation. Current targeted strategies typically involve invasive neurosurgery, whereas systemic approaches produce the undesirable outcome of non-specific protein delivery to the entire brain, rather than solely to the injury site. As a potential way to address this, we developed a protein delivery system modeled after the endogenous immune cell response to brain injury. Using ex-vivo-engineered dendritic cells (DCs, we find that these cells can transiently home to brain injury in a rat model of stroke with both temporal and spatial selectivity. We present a standardized method to derive injury-responsive DCs from bone marrow and show that injury targeting is dependent on culture conditions that maintain an immature DC phenotype. Further, we find evidence that when loaded with therapeutic cargo, cultured DCs can suppress initial neuron death caused by an ischemic injury. These results demonstrate a non-invasive method to target ischemic brain injury and may ultimately provide a way to selectively deliver therapeutic compounds to the injured brain.

  3. Genomic response to Wnt signalling is highly context-dependent - Evidence from DNA microarray and chromatin immunoprecipitation screens of Wnt/TCF targets

    Energy Technology Data Exchange (ETDEWEB)

    Railo, Antti [Oulu Centre for Cell Matrix Research, Biocenter Oulu, Laboratory of Developmental Biology and Department of Medical Biochemistry and Molecular Biology, FIN-90014, University of Oulu, P. O. Box 5000 (Finland); Pajunen, Antti [Department of Biochemistry, University of Oulu (Finland); Itaeranta, Petri; Naillat, Florence; Vuoristo, Jussi; Kilpelaeinen, Pekka [Oulu Centre for Cell Matrix Research, Biocenter Oulu, Laboratory of Developmental Biology and Department of Medical Biochemistry and Molecular Biology, FIN-90014, University of Oulu, P. O. Box 5000 (Finland); Vainio, Seppo, E-mail: Seppo.Vainio@oulu.fi [Oulu Centre for Cell Matrix Research, Biocenter Oulu, Laboratory of Developmental Biology and Department of Medical Biochemistry and Molecular Biology, FIN-90014, University of Oulu, P. O. Box 5000 (Finland)

    2009-10-01

    Wnt proteins are important regulators of embryonic development, and dysregulated Wnt signalling is involved in the oncogenesis of several human cancers. Our knowledge of the downstream target genes is limited, however. We used a chromatin immunoprecipitation-based assay to isolate and characterize the actual gene segments through which Wnt-activatable transcription factors, TCFs, regulate transcription and an Affymetrix microarray analysis to study the global transcriptional response to the Wnt3a ligand. The anti-{beta}-catenin immunoprecipitation of DNA-protein complexes from mouse NIH3T3 fibroblasts expressing a fusion protein of {beta}-catenin and TCF7 resulted in the identification of 92 genes as putative TCF targets. GeneChip assays of gene expression performed on NIH3T3 cells and the rat pheochromocytoma cell line PC12 revealed 355 genes in NIH3T3 and 129 genes in the PC12 cells with marked changes in expression after Wnt3a stimulus. Only 2 Wnt-regulated genes were shared by both cell lines. Surprisingly, Disabled-2 was the only gene identified by the chromatin immunoprecipitation approach that displayed a marked change in expression in the GeneChip assay. Taken together, our approaches give an insight into the complex context-dependent nature of Wnt pathway transcriptional responses and identify Disabled-2 as a potential new direct target for Wnt signalling.

  4. Immune Response to Recombinant Adenovirus in Humans: Capsid Components from Viral Input Are Targets for Vector-Specific Cytotoxic T Lymphocytes

    Science.gov (United States)

    Molinier-Frenkel, Valérie; Gahery-Segard, Hanne; Mehtali, Majid; Le Boulaire, Christophe; Ribault, Sébastien; Boulanger, Pierre; Tursz, Thomas; Guillet, Jean-Gérard; Farace, Françoise

    2000-01-01

    We previously demonstrated that a single injection of 109 PFU of recombinant adenovirus into patients induces strong vector-specific immune responses (H. Gahéry-Ségard, V. Molinier-Frenkel, C. Le Boulaire, P. Saulnier, P. Opolon, R. Lengagne, E. Gautier, A. Le Cesne, L. Zitvogel, A. Venet, C. Schatz, M. Courtney, T. Le Chevalier, T. Tursz, J.-G. Guillet, and F. Farace, J. Clin. Investig. 100:2218–2226, 1997). In the present study we analyzed the mechanism of vector recognition by cytotoxic T lymphocytes (CTL). CD8+ CTL lines were derived from two patients and maintained in long-term cultures. Target cell infections with E1-deleted and E1-plus E2-deleted adenoviruses, as well as transcription-blocking experiments with actinomycin D, revealed that host T-cell recognition did not require viral gene transcription. Target cells treated with brefeldin A were not lysed, indicating that viral input protein-derived peptides are associated with HLA class I molecules. Using recombinant capsid component-loaded targets, we observed that the three major proteins could be recognized. These results raise the question of the use of multideleted adenoviruses for gene therapy in the quest to diminish antivector CTL responses. PMID:10906225

  5. Thermodynamics. Using Affinities to define reversible processes

    CERN Document Server

    Ritacco, Hernán A

    2016-01-01

    In this article a definition of reversible processes in terms of differences in intensive Thermodynamics properties (Affinities) is proposed. This definition makes it possible to both define reversible processes before introducing the concept of entropy and avoid the circularity problem that follows from the Clausius definition of entropy changes. The convenience of this new definition compared to those commonly found in textbooks is demonstrated with examples.

  6. AFFINE TRANSFORMATION IN RANDOM ITERATED FUNCTION SYSTEMS

    Institute of Scientific and Technical Information of China (English)

    熊勇; 史定华

    2001-01-01

    Random iterated function systems (IFSs) is discussed, which is one of the methods for fractal drawing. A certain figure can be reconstructed by a random IFS. One approach is presented to determine a new random IFS, that the figure reconstructed by the new random IFS is the image of the origin figure reconstructed by old IFS under a given affine transformation. Two particular examples are used to show this approach.

  7. Microarray analysis of tomato's early and late wound response reveals new regulatory targets for Leucine aminopeptidase A.

    Directory of Open Access Journals (Sweden)

    Melissa A Scranton

    Full Text Available Wounding due to mechanical injury or insect feeding causes a wide array of damage to plant cells including cell disruption, desiccation, metabolite oxidation, and disruption of primary metabolism. In response, plants regulate a variety of genes and metabolic pathways to cope with injury. Tomato (Solanum lycopersicum is a model for wound signaling but few studies have examined the comprehensive gene expression profiles in response to injury. A cross-species microarray approach using the TIGR potato 10-K cDNA array was analyzed for large-scale temporal (early and late and spatial (locally and systemically responses to mechanical wounding in tomato leaves. These analyses demonstrated that tomato regulates many primary and secondary metabolic pathways and this regulation is dependent on both timing and location. To determine if LAP-A, a known modulator of wound signaling, influences gene expression beyond the core of late wound-response genes, changes in RNAs from healthy and wounded Leucine aminopeptidase A-silenced (LapA-SI and wild-type (WT leaves were examined. While most of the changes in gene expression after wounding in LapA-SI leaves were similar to WT, overall responses were delayed in the LapA-SI leaves. Moreover, two pathogenesis-related 1 (PR-1c and PR-1a2 and two dehydrin (TAS14 and Dhn3 genes were negatively regulated by LAP-A. Collectively, this study has shown that tomato wound responses are complex and that LAP-A's role in modulation of wound responses extends beyond the well described late-wound gene core.

  8. A MEMS Dielectric Affinity Glucose Biosensor.

    Science.gov (United States)

    Huang, Xian; Li, Siqi; Davis, Erin; Li, Dachao; Wang, Qian; Lin, Qiao

    2013-06-20

    Continuous glucose monitoring (CGM) sensors based on affinity detection are desirable for long-term and stable glucose management. However, most affinity sensors contain mechanical moving structures and complex design in sensor actuation and signal readout, limiting their reliability in subcutaneously implantable glucose detection. We have previously demonstrated a proof-of-concept dielectric glucose sensor that measured pre-mixed glucose-sensitive polymer solutions at various glucose concentrations. This sensor features simplicity in sensor design, and possesses high specificity and accuracy in glucose detection. However, lack of glucose diffusion passage, this device is unable to fulfill real-time in-vivo monitoring. As a major improvement to this device, we present in this paper a fully implantable MEMS dielectric affinity glucose biosensor that contains a perforated electrode embedded in a suspended diaphragm. This capacitive-based sensor contains no moving parts, and enables glucose diffusion and real-time monitoring. The experimental results indicate that this sensor can detect glucose solutions at physiological concentrations and possesses good reversibility and reliability. This sensor has a time constant to glucose concentration change at approximately 3 min, which is comparable to commercial systems. The sensor has potential applications in fully implantable CGM that require excellent long-term stability and reliability. PMID:24511215

  9. On constructing purely affine theories with matter

    CERN Document Server

    Cervantes-Cota, Jorge L

    2016-01-01

    We explore ways to obtain the very existence of a space-time metric from an action principle that does not refer to it a priori. Although there are reasons to believe that only a non-local theory can viably achieve this goal, we investigate here local theories that start with Schroedinger's purely affine theory [21], where he gave reasons to set the metric proportional to the Ricci curvature aposteriori. When we leave the context of unified field theory, and we couple the non-gravitational matter using some weak equivalence principle, we can show that the propagation of shock waves does not define a lightcone when the purely affine theory is local and avoids the explicit use of the Ricci tensor in realizing the weak equivalence principle. When the Ricci tensor is substituted for the metric, the equations seem to have only a very limited set of solutions. This backs the conviction that viable purely affine theories have to be non-local.

  10. On constructing purely affine theories with matter

    Science.gov (United States)

    Cervantes-Cota, Jorge L.; Liebscher, D.-E.

    2016-08-01

    We explore ways to obtain the very existence of a space-time metric from an action principle that does not refer to it a priori. Although there are reasons to believe that only a non-local theory can viably achieve this goal, we investigate here local theories that start with Schrödinger's purely affine theory (Schrödinger in Space-time structure. Cambridge UP, Cambridge, 1950), where he gave reasons to set the metric proportional to the Ricci curvature aposteriori. When we leave the context of unified field theory, and we couple the non-gravitational matter using some weak equivalence principle, we can show that the propagation of shock waves does not define a lightcone when the purely affine theory is local and avoids the explicit use of the Ricci tensor in realizing the weak equivalence principle. When the Ricci tensor is substituted for the metric, the equations seem to have only a very limited set of solutions. This backs the conviction that viable purely affine theories have to be non-local.

  11. Increasing the molecular contacts between maurotoxin and Kv1.2 channel augments ligand affinity.

    Science.gov (United States)

    M'Barek, Sarrah; Chagot, Benjamin; Andreotti, Nicolas; Visan, Violeta; Mansuelle, Pascal; Grissmer, Stephan; Marrakchi, Mohamed; El Ayeb, Mohamed; Sampieri, François; Darbon, Hervé; Fajloun, Ziad; De Waard, Michel; Sabatier, Jean-Marc

    2005-08-15

    Scorpion toxins interact with their target ion channels through multiple molecular contacts. Because a "gain of function" approach has never been described to evaluate the importance of the molecular contacts in defining toxin affinity, we experimentally examined whether increasing the molecular contacts between a toxin and an ion channel directly impacts toxin affinity. For this purpose, we focused on two scorpion peptides, the well-characterized maurotoxin with its variant Pi1-like disulfide bridging (MTX(Pi1)), used as a molecular template, and butantoxin (BuTX), used as an N-terminal domain provider. BuTX is found to be 60-fold less potent than MTX(Pi1) in blocking Kv1.2 (IC(50) values of 165 nM for BuTX versus 2.8 nM for MTX(Pi1)). Removal of its N-terminal domain (nine residues) further decreases BuTX affinity for Kv1.2 by 5.6-fold, which is in agreement with docking simulation data showing the importance of this domain in BuTX-Kv1.2 interaction. Transfer of the BuTX N-terminal domain to MTX(Pi1) results in a chimera with five disulfide bridges (BuTX-MTX(Pi1)) that exhibits 22-fold greater affinity for Kv1.2 than MTX(Pi1) itself, in spite of the lower affinity of BuTX as compared to MTX(Pi1). Docking experiments performed with the 3-D structure of BuTX-MTX(Pi1) in solution, as solved by (1)H-NMR, reveal that the N-terminal domain of BuTX participates in the increased affinity for Kv1.2 through additional molecular contacts. Altogether, the data indicate that acting on molecular contacts between a toxin and a channel is an efficient strategy to modulate toxin affinity.

  12. BCL-2 family proteins as 5-Azacytidine-sensitizing targets and determinants of response in myeloid malignancies

    OpenAIRE

    Bogenberger, J M; Kornblau, S. M.; Pierceall, W E; Lena, R.; Chow, D.; Shi, C-X; Mantei, J; Ahmann, G; Gonzales, I M; A. Choudhary; R. Valdez; Camoriano, J; Fauble, V; Tiedemann, R E; Qiu, Y H

    2014-01-01

    Synergistic molecular vulnerabilities enhancing hypomethylating agents in myeloid malignancies have remained elusive. RNA-interference drug modifier screens identified antiapoptotic BCL-2 family members as potent 5-Azacytidine-sensitizing targets. In further dissecting BCL-XL, BCL-2 and MCL-1 contribution to 5-Azacytidine activity, siRNA silencing of BCL-XL and MCL-1, but not BCL-2, exhibited variable synergy with 5-Azacytidine in vitro. The BCL-XL, BCL-2 and BCL-w inhibitor ABT-737 sensitize...

  13. MicroRNA-155 Targets SMAD2 and Modulates the Response of Macrophages to Transforming Growth Factor-β*

    OpenAIRE

    Louafi, Fethi; Martinez-Nunez, Rocio T.; Sanchez-Elsner, Tilman

    2010-01-01

    Transforming growth factor-beta (TGF-β) is a pleiotropic cytokine with important effects on processes such as fibrosis, angiogenesis, and immunosupression. Using bioinformatics, we identified SMAD2, one of the mediators of TGF-β signaling, as a predicted target for a microRNA, microRNA-155 (miR-155). MicroRNAs are a class of small non-coding RNAs that have emerged as an important class of gene expression regulators. miR-155 has been found to be involved in the regulation of the immune respons...

  14. Genomic Copy Number Dictates a Gene-Independent Cell Response to CRISPR/Cas9 Targeting | Office of Cancer Genomics

    Science.gov (United States)

    The CRISPR/Cas9 system enables genome editing and somatic cell genetic screens in mammalian cells. We performed genome-scale loss-of-function screens in 33 cancer cell lines to identify genes essential for proliferation/survival and found a strong correlation between increased gene copy number and decreased cell viability after genome editing. Within regions of copy-number gain, CRISPR/Cas9 targeting of both expressed and unexpressed genes, as well as intergenic loci, led to significantly decreased cell proliferation through induction of a G2 cell-cycle arrest.

  15. Bacterial rRNA-Targeted Reverse Transcription-PCR Used To Identify Pathogens Responsible for Fever with Neutropenia▿

    OpenAIRE

    Sakaguchi, Sachi; Saito, Masahiro; Tsuji, Hirokazu; Asahara, Takashi; Takata, Oto; Fujimura, Junya; NAGATA, Satoru; Nomoto, Koji; Shimizu, Toshiaki

    2010-01-01

    The purpose of this study was to evaluate the clinical utility of bacterial rRNA-targeted reverse transcription-quantitative PCR (BrRNA RT-qPCR) assays for identifying the bacterial pathogens that cause fever with neutropenia in pediatric cancer patients, by comparing the bacterial detection rate of this technique with that of blood culture. One milliliter of blood was collected from pediatric patients who developed fever with neutropenia following cancer chemotherapy. BrRNA RT-qPCR was perfo...

  16. A graphitic hollow carbon nitride nanosphere as a novel photochemical internalization agent for targeted and stimuli-responsive cancer therapy

    Science.gov (United States)

    Liu, Chaoqun; Chen, Zhaowei; Wang, Zhenzhen; Li, Wei; Ju, Enguo; Yan, Zhengqing; Liu, Zhen; Ren, Jinsong; Qu, Xiaogang

    2016-06-01

    As a novel technique, photochemical internalization (PCI) has been employed as a new approach to overcome endo/lysosomal restriction, which is one of the main difficulties in both drug and gene delivery. However, the complicated synthesis procedure (usually requiring the self-assembly of polymers, photosensitizers and cargos) and payload specificity greatly limit its further application. In this paper, we employ a highly fluorescent graphitic hollow carbon nitride nanosphere (GHCNS) to simultaneously serve as a PCI photosensitizer, an imaging agent and a drug carrier. The surface modification of GHCNS with multifunctional polysaccharide hyaluronic acid (HA) endows the system with colloidal stability, biocompatibility and cancer cell targeting ability. After CD44 receptor-mediated endocytosis, the nanosystem is embedded in endo/lysosomal vesicles and HA could be specially degraded by hyaluronidase (Hyal), inducing open pores. In the following, with visible light illumination, GHCNS could produce ROS that effectively induced lipid peroxidation and caused endo/lysosomal membrane break, accelerating the cytoplasmic release of the drug in the targeted and irradiated cells. As a result, significantly increased therapeutic potency and specificity against cancer cells could be achieved.As a novel technique, photochemical internalization (PCI) has been employed as a new approach to overcome endo/lysosomal restriction, which is one of the main difficulties in both drug and gene delivery. However, the complicated synthesis procedure (usually requiring the self-assembly of polymers, photosensitizers and cargos) and payload specificity greatly limit its further application. In this paper, we employ a highly fluorescent graphitic hollow carbon nitride nanosphere (GHCNS) to simultaneously serve as a PCI photosensitizer, an imaging agent and a drug carrier. The surface modification of GHCNS with multifunctional polysaccharide hyaluronic acid (HA) endows the system with colloidal

  17. In vivo transcription of a progesterone-responsive gene is specifically inhibited by a triplex-forming oligonucleotide.

    OpenAIRE

    Ing, N H; Beekman, J M; Kessler, D J; Murphy, M.; Jayaraman, K; Zendegui, J G; Hogan, M E; O'Malley, B W; Tsai, M J

    1993-01-01

    Oligonucleotides provide novel reagents for inhibition of gene expression because of their high affinity binding to specific nucleotide sequences. We describe a 38 base, single-stranded DNA that forms a triple helix or 'triplex' on progesterone response elements of a target gene. This triplex-forming oligonucleotide binds with a Kd = 100 nM at 37 degrees C and physiological pH, and blocks binding of progesterone receptors to the target. Furthermore, it completely inhibited progesterone recept...

  18. Experimental investigation of streamer affinity for dielectric surfaces

    NARCIS (Netherlands)

    Trienekens, D.J.M.; Nijdam, S.; Akkermans, G.; Plompen, I.; Christen, T.; Ebert, U.

    2015-01-01

    We have experimentally investigated the affinity of streamers for dielectric surfaces using stroboscopic imaging and stereo photography. Affinity of streamers for dielectric surfaces was found to depend on a wide set of parameters, including pressure, voltage, dielectric material and di

  19. Quelques remarques sur la notion de modification affine

    OpenAIRE

    Dubouloz, Adrien

    2005-01-01

    in french We construct a global counterpart to the notion of affine modification due to Kaliman and Zaidenberg. This leads to a simple explicit description of the structure of birational affine morphisms between arbitrary quasi-projective varieties.

  20. Regulation of the heat stress response in Arabidopsis by MPK6-targeted phosphorylation of the heat stress factor HsfA2

    Directory of Open Access Journals (Sweden)

    Alexandre Evrard

    2013-04-01

    Full Text Available So far little is known on the functional role of phosphorylation in the heat stress response of plants. Here we present evidence that heat stress activates the Arabidopsis mitogen-activated protein kinase MPK6. In vitro and in vivo evidence is provided that MPK6 specifically targets the major heat stress transcription factor HsfA2. Activation of MPK6 results in complex formation with HsfA2. MPK6 phosphorylates HsfA2 on T249 and changes its intracellular localisation. Protein kinase and phosphatase inhibitor studies indicate that HsfA2 protein stability is regulated in a phosphorylation-dependent manner, but this mechanism is independent of MPK6. Overall, our data show that heat stress-induced targeting of HsfA2 by MPK6 participates in the complex regulatory mechanism how plants respond to heat stress.

  1. EGFRvIII deletion mutations in pediatric high-grade glioma and response to targeted therapy in pediatric glioma cell lines

    DEFF Research Database (Denmark)

    Bax, Dorine A; Gaspar, Nathalie; Little, Suzanne E;

    2009-01-01

    to erlotinib despite expressing wild-type PTEN. Phosphorylated receptor tyrosine kinase profiling showed a specific activation of platelet-derived growth factor receptor alpha/beta in EGFRvIII-transduced pediatric glioblastoma cells, and targeted coinhibition with erlotinib and imatinib leads to enhanced......PURPOSE: The epidermal growth factor receptor (EGFR) is amplified and overexpressed in adult glioblastoma, with response to targeted inhibition dependent on the underlying biology of the disease. EGFR has thus far been considered to play a less important role in pediatric glioma, although extensive...... of pediatric glioma cell line models to the small-molecule EGFR inhibitor erlotinib. RESULTS: Amplification was detected in 11% of cases, with corresponding overexpression of the receptor. No kinase or extracellular domain mutations were observed; however, 6 of 35 (17%) cases harbored the EGFRvIII deletion...

  2. Computational protocol for predicting the binding affinities of zinc containing metalloprotein-ligand complexes.

    Science.gov (United States)

    Jain, Tarun; Jayaram, B

    2007-06-01

    Zinc is one of the most important metal ions found in proteins performing specific functions associated with life processes. Coordination geometry of the zinc ion in the active site of the metalloprotein-ligand complexes poses a challenge in determining ligand binding affinities accurately in structure-based drug design. We report here an all atom force field based computational protocol for estimating rapidly the binding affinities of zinc containing metalloprotein-ligand complexes, considering electrostatics, van der Waals, hydrophobicity, and loss in conformational entropy of protein side chains upon ligand binding along with a nonbonded approach to model the interactions of the zinc ion with all the other atoms of the complex. We examined the sensitivity of the binding affinity predictions to the choice of Lennard-Jones parameters, partial atomic charges, and dielectric treatments adopted for system preparation and scoring. The highest correlation obtained was R2 = 0.77 (r = 0.88) for the predicted binding affinity against the experiment on a heterogenous dataset of 90 zinc containing metalloprotein-ligand complexes consisting of five unique protein targets. Model validation and parameter analysis studies underscore the robustness and predictive ability of the scoring function. The high correlation obtained suggests the potential applicability of the methodology in designing novel ligands for zinc-metalloproteins. The scoring function has been web enabled for free access at www.scfbio-iitd.res.in/software/drugdesign/bapplz.jsp as BAPPL-Z server (Binding Affinity Prediction of Protein-Ligand complexes containing Zinc metal ions).

  3. Parameterization of an effective potential for protein-ligand binding from host-guest affinity data.

    Science.gov (United States)

    Wickstrom, Lauren; Deng, Nanjie; He, Peng; Mentes, Ahmet; Nguyen, Crystal; Gilson, Michael K; Kurtzman, Tom; Gallicchio, Emilio; Levy, Ronald M

    2016-01-01

    Force field accuracy is still one of the "stalemates" in biomolecular modeling. Model systems with high quality experimental data are valuable instruments for the validation and improvement of effective potentials. With respect to protein-ligand binding, organic host-guest complexes have long served as models for both experimental and computational studies because of the abundance of binding affinity data available for such systems. Binding affinity data collected for cyclodextrin (CD) inclusion complexes, a popular model for molecular recognition, is potentially a more reliable resource for tuning energy parameters than hydration free energy measurements. Convergence of binding free energy calculations on CD host-guest systems can also be obtained rapidly, thus offering the opportunity to assess the robustness of these parameters. In this work, we demonstrate how implicit solvent parameters can be developed using binding affinity experimental data and the binding energy distribution analysis method (BEDAM) and validated using the Grid Inhomogeneous Solvation Theory analysis. These new solvation parameters were used to study protein-ligand binding in two drug targets against the HIV-1 virus and improved the agreement between the calculated and the experimental binding affinities. This work illustrates how benchmark sets of high quality experimental binding affinity data and physics-based binding free energy models can be used to evaluate and optimize force fields for protein-ligand systems. Copyright © 2015 John Wiley & Sons, Ltd. PMID:26256816

  4. Applications of Aptamers in Targeted Imaging: State of the Art

    OpenAIRE

    Dougherty, Casey A.; Cai, Weibo; Hong, Hao

    2015-01-01

    Aptamers are single-stranded oligonucleotides with high affinity and specificity to the target molecules or cells, thus they can serve as an important category of molecular targeting ligand. Since their discove1y, aptamers have been rapidly translated into clinical practice. The strong target affinity/selectivity, cost-effectivity, chemical versatility and safety of aptamers are superior to traditional peptides- or proteins-based ligands which make them unique choices for molecular imaging. T...

  5. Immunological Cross-Reactivity between Malaria Vaccine Target Antigen P48/45 in Plasmodium vivax and P. falciparum and Cross-Boosting of Immune Responses.

    Science.gov (United States)

    Cao, Yi; Bansal, Geetha P; Merino, Kristen; Kumar, Nirbhay

    2016-01-01

    In general, malaria immunity has been suggested to be species specific with very little, if any, known cross-reactivity between Plasmodium vivax and P. falciparum, both of which are responsible for >90% of human malaria, and co-endemic in many countries. It is therefore believed that species-specific immunity may be needed to target different species of Plasmodium. Pfs48/45 and Pvs48/45 are well established targets in the sexual stages of the malaria parasites, and are being pursued for the development of transmission blocking vaccines. Comparison of their sequences reveals 61% and 55% identity at the DNA and protein level, respectively raising the possibility that these two target antigens might share cross-reacting epitopes. Having succeeded in expressing recombinant Pfs48/45 and Pvs48/45 proteins, we hypothesized that these proteins will not only exhibit immunological cross-reactivity but also cross-boost immune responses. Mice were immunized with purified recombinant proteins using CFA, Montanide ISA-51 and alum as adjuvants, and the sera were analyzed by ELISA, Western blotting and indirect fixed and live IFA to address the hypothesis. Our studies revealed that Pvs48/45-immune sera showed strong cross-reactivity to full length Pfs48/45 protein, and the majority of this cross reactivity was in the amino-terminal and carboxyl-terminal sub-fragments of Pfs48/45. In cross-boosting experiments Pfs48/45 and Pvs48/45 antigens were able to cross-boost each other in mouse immunization studies. Additionally we also noticed an effect of adjuvants in the overall magnitude of observed cross-reactivity. These studies may have significant implications for immunity targeting transmission of both the species of malaria parasites. PMID:27438603

  6. Immunological Cross-Reactivity between Malaria Vaccine Target Antigen P48/45 in Plasmodium vivax and P. falciparum and Cross-Boosting of Immune Responses.

    Directory of Open Access Journals (Sweden)

    Yi Cao

    Full Text Available In general, malaria immunity has been suggested to be species specific with very little, if any, known cross-reactivity between Plasmodium vivax and P. falciparum, both of which are responsible for >90% of human malaria, and co-endemic in many countries. It is therefore believed that species-specific immunity may be needed to target different species of Plasmodium. Pfs48/45 and Pvs48/45 are well established targets in the sexual stages of the malaria parasites, and are being pursued for the development of transmission blocking vaccines. Comparison of their sequences reveals 61% and 55% identity at the DNA and protein level, respectively raising the possibility that these two target antigens might share cross-reacting epitopes. Having succeeded in expressing recombinant Pfs48/45 and Pvs48/45 proteins, we hypothesized that these proteins will not only exhibit immunological cross-reactivity but also cross-boost immune responses. Mice were immunized with purified recombinant proteins using CFA, Montanide ISA-51 and alum as adjuvants, and the sera were analyzed by ELISA, Western blotting and indirect fixed and live IFA to address the hypothesis. Our studies revealed that Pvs48/45-immune sera showed strong cross-reactivity to full length Pfs48/45 protein, and the majority of this cross reactivity was in the amino-terminal and carboxyl-terminal sub-fragments of Pfs48/45. In cross-boosting experiments Pfs48/45 and Pvs48/45 antigens were able to cross-boost each other in mouse immunization studies. Additionally we also noticed an effect of adjuvants in the overall magnitude of observed cross-reactivity. These studies may have significant implications for immunity targeting transmission of both the species of malaria parasites.

  7. Immunological Cross-Reactivity between Malaria Vaccine Target Antigen P48/45 in Plasmodium vivax and P. falciparum and Cross–Boosting of Immune Responses

    Science.gov (United States)

    Cao, Yi; Bansal, Geetha P.; Merino, Kristen; Kumar, Nirbhay

    2016-01-01

    In general, malaria immunity has been suggested to be species specific with very little, if any, known cross-reactivity between Plasmodium vivax and P. falciparum, both of which are responsible for >90% of human malaria, and co-endemic in many countries. It is therefore believed that species-specific immunity may be needed to target different species of Plasmodium. Pfs48/45 and Pvs48/45 are well established targets in the sexual stages of the malaria parasites, and are being pursued for the development of transmission blocking vaccines. Comparison of their sequences reveals 61% and 55% identity at the DNA and protein level, respectively raising the possibility that these two target antigens might share cross-reacting epitopes. Having succeeded in expressing recombinant Pfs48/45 and Pvs48/45 proteins, we hypothesized that these proteins will not only exhibit immunological cross–reactivity but also cross-boost immune responses. Mice were immunized with purified recombinant proteins using CFA, Montanide ISA-51 and alum as adjuvants, and the sera were analyzed by ELISA, Western blotting and indirect fixed and live IFA to address the hypothesis. Our studies revealed that Pvs48/45-immune sera showed strong cross-reactivity to full length Pfs48/45 protein, and the majority of this cross reactivity was in the amino-terminal and carboxyl-terminal sub-fragments of Pfs48/45. In cross-boosting experiments Pfs48/45 and Pvs48/45 antigens were able to cross-boost each other in mouse immunization studies. Additionally we also noticed an effect of adjuvants in the overall magnitude of observed cross-reactivity. These studies may have significant implications for immunity targeting transmission of both the species of malaria parasites. PMID:27438603

  8. Simulation of Thermal Responses of 125TeO2 Solid Target to Energetic Proton Bombardment from Cyclotron When Fabricating 124I Nuclear Medicine

    Science.gov (United States)

    Peir, Jinn-Jer; Liang, Jenq-Horng; Duh, Ting-Shieh

    With nuclear medicine receiving greater attention due to its unique characteristics in both diagnostics and therapeutics during recent decades, finding a highly controllable fabrication method becomes more urgent. The radioisotope 124I (T1/2=4.18d Eβ+=2.13MeV Iβ+=25%) has gained plentiful interests in the medical usages such as functioning imaging of cell proliferation in brain tumors using [124I]iododeoxyuridine (IUdR), imaging of immunoreactions in tumors using 124I-labelled monoclonal antibodies, the in-vivo imaging of 124I-labelled tyrosine derivatives, and the classical imaging of thyroid diseases with 124I, among others. Furthermore, it is because that thermal response of target during the fabrication process may affect the production of 124I to some extent and needs thorough investigations. Hence, the compact cyclotron located in the Institute of Nuclear Energy Research was employed in this study to generate 20MeV protons to irradiate TeO2 solid targets in which the radioisotopes 124I were produced through the 125Te(p, 2n)124I nuclear reaction. In addition, the widely-used ANSYS computer code was adopted to theoretically analyze thermal responses of TeO2 to irradiation cases with variations in ion beam current and its thermal conductivity. The results indicate that TeO2 temperature is strongly dependent on its thermal conductivity and ion beam current. In particular, TeO2 surface temperature is extremely sensitive to the air-gap size between TeO2 and target holder. Thus the target holder is suggested to be re-designed in order to prevent TeO2 from melting and a high efficiency production of radioisotopes 124I for nuclear medical diagnostics can be successfully achieved.

  9. Novel Nuclear Factor-KappaB Targeting Peptide Suppresses β-Amyloid Induced Inflammatory and Apoptotic Responses in Neuronal Cells

    Science.gov (United States)

    Srinivasan, Mythily; Bayon, Baindu; Chopra, Nipun; Lahiri, Debomoy K.

    2016-01-01

    In the central nervous system (CNS), activation of the transcription factor nuclear factor-kappa B (NF-κβ) is associated with both neuronal survival and increased vulnerability to apoptosis. The mechanisms underlying these dichotomous effects are attributed to the composition of NF-κΒ dimers. In Alzheimer’s disease (AD), β-amyloid (Aβ) and other aggregates upregulate activation of p65:p50 dimers in CNS cells and enhance transactivation of pathological mediators that cause neuroinflammation and neurodegeneration. Hence selective targeting of activated p65 is an attractive therapeutic strategy for AD. Here we report the design, structural and functional characterization of peptide analogs of a p65 interacting protein, the glucocorticoid induced leucine zipper (GILZ). By virtue of binding the transactivation domain of p65 exposed after release from the inhibitory IκΒ proteins in activated cells, the GILZ analogs can act as highly selective inhibitors of activated p65 with minimal potential for off-target effects. PMID:27764084

  10. Fractal Homeomorphism for Bi-affine Iterated Function Systems

    CERN Document Server

    Barnsley, Michael

    2011-01-01

    The paper concerns fractal homeomorphism between the attractors of two bi-affine iterated function systems. After a general discussion of bi-affine functions, conditions are provided under which a bi-affine iterated function system is contractive, thus guaranteeing an attractor. After a general discussion of fractal homeomorphism, fractal homeomorphisms are constructed for a specific type of bi-affine iterated function system.

  11. The purification of affinity-labelled active-site peptides

    International Nuclear Information System (INIS)

    The isolation of the labelled peptide from the protein digest, following the affinity labelling of the active sites of enzymes or antibodies, is described. Single-step affinity chromatography utilises the affinity of the native enzymes or antibody for the ligand used to label the same protein. The labelled peptide is the only one in the digest that displays affinity for the immobilised protein and can be released with eluants that dissociate the protein-ligand complex. (Auth.)

  12. Structural and Nonstructural Viral Proteins Are Targets of T-Helper Immune Response against Human Respiratory Syncytial Virus.

    Science.gov (United States)

    Lorente, Elena; Barriga, Alejandro; Barnea, Eilon; Mir, Carmen; Gebe, John A; Admon, Arie; López, Daniel

    2016-06-01

    Proper antiviral humoral and cellular immune responses require previous recognition of viral antigenic peptides that are bound to HLA class II molecules, which are exposed on the surface of antigen-presenting cells. The helper immune response is critical for the control and the clearance of human respiratory syncytial virus (HRSV) infection, a virus with severe health risk in infected pediatric, immunocompromised, and elderly populations. In this study, using a mass spectrometry analysis of complex HLA class II-bound peptide pools that were isolated from large amounts of HRSV-infected cells, 19 naturally processed HLA-DR ligands, most of them included in a complex nested set of peptides, were identified. Both the immunoprevalence and the immunodominance of the HLA class II response to HRSV were focused on one nonstructural (NS1) and two structural (matrix and mainly fusion) proteins of the infective virus. These findings have clear implications for analysis of the helper immune response as well as for antiviral vaccine design. PMID:27090790

  13. UBXN1 Interferes with Rig-I-like Receptor-Mediated Antiviral Immune Response by Targeting MAVS

    Directory of Open Access Journals (Sweden)

    Penghua Wang

    2013-04-01

    Full Text Available RNA viruses are sensed by RIG-I-like receptors (RLRs, which signal through a mitochondria-associated adaptor molecule, MAVS, resulting in systemic antiviral immune responses. Although RLR signaling is essential for limiting RNA virus replication, it must be stringently controlled to prevent damage from inflammation. We demonstrate here that among all tested UBX-domain-containing protein family members, UBXN1 exhibits the strongest inhibitory effect on RNA-virus-induced type I interferon response. UBXN1 potently inhibits RLR- and MAVS-induced, but not TLR3-, TLR4-, or DNA-virus-induced innate immune responses. Depletion of UBXN1 enhances virus-induced innate immune responses, including those resulting from RNA viruses such as vesicular stomatitis, Sendai, West Nile, and dengue virus infection, repressing viral replication. Following viral infection, UBXN1 is induced, binds to MAVS, interferes with intracellular MAVS oligomerization, and disrupts the MAVS/TRAF3/TRAF6 signalosome. These findings underscore a critical role of UBXN1 in the modulation of a major antiviral signaling pathway.

  14. Target Practice: Reader Response Theory and Teachers' Interpretations of Students' SAT 10 Scores in Data-Based Professional Development

    Science.gov (United States)

    Atkinson, Becky M.

    2012-01-01

    The study reported in this article examines how teachers read and respond to their students' Stanford Achievement Test 10 (SAT 10) scores with the goal of investigating the assumption that data-based teaching practice is more "objective" and less susceptible to divergent teacher interpretation. The study uses reader response theory to frame…

  15. Targeting Werner syndrome protein sensitizes U-2 OS osteosarcoma cells to selenium-induced DNA damage response and necrotic death

    DEFF Research Database (Denmark)

    Cheng, Wen-Hsing; Wu, Ryan T Y; Wu, Min;

    2012-01-01

    Mutations in the Werner syndrome protein (WRN), a caretaker of the genome, result in Werner syndrome, which is characterized by premature aging phenotypes and cancer predisposition. Methylseleninic acid (MSeA) can activate DNA damage responses and is a superior compound to suppress tumorigenesis ...

  16. Doxorubicin loaded dual pH- and thermo-responsive magnetic nanocarrier for combined magnetic hyperthermia and targeted controlled drug delivery applications

    Science.gov (United States)

    Hervault, Aziliz; Dunn, Alexander E.; Lim, May; Boyer, Cyrille; Mott, Derrick; Maenosono, Shinya; Thanh, Nguyen T. K.

    2016-06-01

    Magnetic nanocarriers have attracted increasing attention for multimodal cancer therapy due to the possibility to deliver heat and drugs locally. The present study reports the development of magnetic nanocomposites (MNCs) made of an iron oxide core and a pH- and thermo-responsive polymer shell, that can be used as both hyperthermic agent and drug carrier. The conjugation of anticancer drug doxorubicin (DOX) to the pH- and thermo-responsive MNCs via acid-cleavable imine linker provides advanced features for the targeted delivery of DOX molecules via the combination of magnetic targeting, and dual pH- and thermo-responsive behaviour which offers spatial and temporal control over the release of DOX. The iron oxide cores exhibit a superparamagnetic behaviour with a saturation magnetization around 70 emu g-1. The MNCs contained 8.1 wt% of polymer and exhibit good heating properties in an alternating magnetic field. The drug release experiments confirmed that only a small amount of DOX was released at room temperature and physiological pH, while the highest drug release of 85.2% was obtained after 48 h at acidic tumour pH under hyperthermia conditions (50 °C). The drug release kinetic followed Korsmeyer-Peppas model and displayed Fickian diffusion mechanism. From the results obtained it can be concluded that this smart magnetic nanocarrier is promising for applications in multi-modal cancer therapy, to target and efficiently deliver heat and drug specifically to the tumour.Magnetic nanocarriers have attracted increasing attention for multimodal cancer therapy due to the possibility to deliver heat and drugs locally. The present study reports the development of magnetic nanocomposites (MNCs) made of an iron oxide core and a pH- and thermo-responsive polymer shell, that can be used as both hyperthermic agent and drug carrier. The conjugation of anticancer drug doxorubicin (DOX) to the pH- and thermo-responsive MNCs via acid-cleavable imine linker provides advanced

  17. The oncogenic MicroRNA Hsa-miR-155-5p targets the transcription factor ELK3 and links it to the hypoxia response.

    Directory of Open Access Journals (Sweden)

    E Douglas Robertson

    Full Text Available The molecular response to hypoxia is a critical cellular process implicated in cancer, and a target for drug development. The activity of the major player, HIF1α, is regulated at different levels by various factors, including the transcription factor ELK3. The molecular mechanisms of this intimate connection remain largely unknown. Whilst investigating global ELK3-chromatin interactions, we uncovered an unexpected connection that involves the microRNA hsa-miR-155-5p, a hypoxia-inducible oncomir that targets HIF1α. One of the ELK3 chromatin binding sites, detected by Chromatin Immuno-Precipitation Sequencing (ChIP-seq of normal Human Umbilical Vein Endothelial Cells (HUVEC, is located at the transcription start site of the MIR155HG genes that expresses hsa-miR-155-5p. We confirmed that ELK3 binds to this promoter by ChIP and quantitative polymerase chain reaction (QPCR. We showed that ELK3 and hsa-miR-155-5p form a double-negative regulatory loop, in that ELK3 depletion induced hsa-miR-155-5p expression and hsa-miR-155-5p expression decreased ELK3 expression at the RNA level through a conserved target sequence in its 3'-UTR. We further showed that the activities of hsa-miR-155-5p and ELK3 are functionally linked. Pathway analysis indicates that both factors are implicated in related processes, including cancer and angiogenesis. Hsa-miR-155-5p expression and ELK3 depletion have similar effects on expression of known ELK3 target genes, and on in-vitro angiogenesis and wound closure. Bioinformatic analysis of cancer RNA-seq data shows that hsa-miR-155-5p and ELK3 expression are significantly anti-correlated, as would be expected from hsa-miR-155-5p targeting ELK3 RNA. Finally, hypoxia (0% oxygen down-regulates ELK3 mRNA in a microRNA and hsa-miR-155-5p dependent manner. These results tie ELK3 into the hypoxia response pathway through an oncogenic microRNA and into a circuit implicated in the dynamics of the hypoxic response. This crosstalk could

  18. The oncogenic MicroRNA Hsa-miR-155-5p targets the transcription factor ELK3 and links it to the hypoxia response.

    Science.gov (United States)

    Robertson, E Douglas; Wasylyk, Christine; Ye, Tao; Jung, Alain C; Wasylyk, Bohdan

    2014-01-01

    The molecular response to hypoxia is a critical cellular process implicated in cancer, and a target for drug development. The activity of the major player, HIF1α, is regulated at different levels by various factors, including the transcription factor ELK3. The molecular mechanisms of this intimate connection remain largely unknown. Whilst investigating global ELK3-chromatin interactions, we uncovered an unexpected connection that involves the microRNA hsa-miR-155-5p, a hypoxia-inducible oncomir that targets HIF1α. One of the ELK3 chromatin binding sites, detected by Chromatin Immuno-Precipitation Sequencing (ChIP-seq) of normal Human Umbilical Vein Endothelial Cells (HUVEC), is located at the transcription start site of the MIR155HG genes that expresses hsa-miR-155-5p. We confirmed that ELK3 binds to this promoter by ChIP and quantitative polymerase chain reaction (QPCR). We showed that ELK3 and hsa-miR-155-5p form a double-negative regulatory loop, in that ELK3 depletion induced hsa-miR-155-5p expression and hsa-miR-155-5p expression decreased ELK3 expression at the RNA level through a conserved target sequence in its 3'-UTR. We further showed that the activities of hsa-miR-155-5p and ELK3 are functionally linked. Pathway analysis indicates that both factors are implicated in related processes, including cancer and angiogenesis. Hsa-miR-155-5p expression and ELK3 depletion have similar effects on expression of known ELK3 target genes, and on in-vitro angiogenesis and wound closure. Bioinformatic analysis of cancer RNA-seq data shows that hsa-miR-155-5p and ELK3 expression are significantly anti-correlated, as would be expected from hsa-miR-155-5p targeting ELK3 RNA. Finally, hypoxia (0% oxygen) down-regulates ELK3 mRNA in a microRNA and hsa-miR-155-5p dependent manner. These results tie ELK3 into the hypoxia response pathway through an oncogenic microRNA and into a circuit implicated in the dynamics of the hypoxic response. This crosstalk could be important for

  19. [Expression analysis of miR164 and its target gene NAC1 in response to low nitrate availability in Betula luminifera].

    Science.gov (United States)

    Jun, Wu; Junhong, Zhang; Menghui, Huang; Minhui, Zhu; Zaikang, Tong

    2016-02-01

    Nitrogen, an essential macronutrient for the growth and development of plants, affects above- ground biomass accumulation dramatically. Thus, it is very important to reveal the molecular mechanisms of how plants resist or adapt to low nitrogen availability. The NAC1(NAM, ATAF, CUC 1) gene, located in the upstream regulatory network, has been reported to resist low nitrogen by regulating expression of key downstream genes and thus root growth in (Populus tremula × alba).In this study, we detected the responses of miR164 and its target gene NAC1 under nitrate-starvation condition using the Betula luminifera somaclones G49-3 as material. The NAC1 gene which contains 1497 bp sequence, encodes 358 amino acids and contains a highly conserved NAM domain at N terminal was cloned by the RACE method. The NAC1 was then validated to be the target gene of miR164 via 5'-RACE, and the cleavage site was between the 10(th) and 11(th) base. The expression patterns of miR164 and its target gene NAC1 were further detected under nitrate-starvation condition through qRT-PCR analysis. The results showed that miR164 expression was repressed by nitrate-starvation at the beginning of the treatment (4 d) and then ascended. However, the expression pattern of miR164 in roots was different from that in shoots and leaves. Moreover, the expression levels of target gene NAC1 and miR164 were negatively correlated. The expression level of miR164 in root was increased while that of NAC1 was decreased under Re treatment, which indicated that miR164 and its target gene NAC1 play a regulatory role in response to low nitrate availability. The findings of our study may help elucidate the molecular mechanisms by which miR164 regulates target gene NAC1 at post-transcriptional level, and provide valuable information for further study of the regulatory roles of miR164-NAC1 under nitrate-starvation condition.

  20. Comparative Analysis and Identification of miRNAs and Their Target Genes Responsive to Salt Stress in Diploid and Tetraploid Paulownia fortunei Seedlings.

    Directory of Open Access Journals (Sweden)

    Guoqiang Fan

    Full Text Available Salt stress is a global environmental problem that affects plant growth and development. Paulownia fortunei is an adaptable and fast-growing deciduous tree native to China that is environmentally and economically important. MicroRNAs (miRNAs play important regulatory roles in growth, development, and stress responses in plants. MiRNAs that respond to biotic stresses have been identified; however, how miRNAs in P. fortunei respond to salt stress has not yet been reported. To identify salt-stress-responsive miRNAs and predict their target genes, four small RNA and four degradome libraries were constructed from NaCl-treated and NaCl-free leaves of P. fortunei seedlings. The results indicated that salt stress had different physiological effects on diploid and tetraploid P. fortunei. We detected 53 conserved miRNAs belonging to 17 miRNA families and 134 novel miRNAs in P. fortunei. Comparing their expression levels in diploid and tetraploid P. fortunei, we found 10 conserved and 10 novel miRNAs that were significantly differentially expressed under salt treatment, among them eight were identified as miRNAs probably associated with higher salt tolerance in tetraploid P. fortunei than in diploid P. fortunei. Gene Ontology and Kyoto Encyclopedia of Genes and Genomes pathway analyses were performed to predict the functions of the target genes of the conserved and novel miRNAs. The expressions of 10 differentially expressed miRNAs were validated by quantitative real-time polymerase chain reaction (qRT-PCR. This is the first report on P. fortunei miRNAs and their target genes under salt stress. The results provided information at the physiological and molecular levels for further research into the response mechanisms of P. fortunei to salt stress.

  1. Comparative Analysis and Identification of miRNAs and Their Target Genes Responsive to Salt Stress in Diploid and Tetraploid Paulownia fortunei Seedlings.

    Science.gov (United States)

    Fan, Guoqiang; Li, Xiaoyu; Deng, Minjie; Zhao, Zhenli; Yang, Lu

    2016-01-01

    Salt stress is a global environmental problem that affects plant growth and development. Paulownia fortunei is an adaptable and fast-growing deciduous tree native to China that is environmentally and economically important. MicroRNAs (miRNAs) play important regulatory roles in growth, development, and stress responses in plants. MiRNAs that respond to biotic stresses have been identified; however, how miRNAs in P. fortunei respond to salt stress has not yet been reported. To identify salt-stress-responsive miRNAs and predict their target genes, four small RNA and four degradome libraries were constructed from NaCl-treated and NaCl-free leaves of P. fortunei seedlings. The results indicated that salt stress had different physiological effects on diploid and tetraploid P. fortunei. We detected 53 conserved miRNAs belonging to 17 miRNA families and 134 novel miRNAs in P. fortunei. Comparing their expression levels in diploid and tetraploid P. fortunei, we found 10 conserved and 10 novel miRNAs that were significantly differentially expressed under salt treatment, among them eight were identified as miRNAs probably associated with higher salt tolerance in tetraploid P. fortunei than in diploid P. fortunei. Gene Ontology and Kyoto Encyclopedia of Genes and Genomes pathway analyses were performed to predict the functions of the target genes of the conserved and novel miRNAs. The expressions of 10 differentially expressed miRNAs were validated by quantitative real-time polymerase chain reaction (qRT-PCR). This is the first report on P. fortunei miRNAs and their target genes under salt stress. The results provided information at the physiological and molecular levels for further research into the response mechanisms of P. fortunei to salt stress. PMID:26894691

  2. Immuno and affinity cytochemical analysis of cell wall composition in the moss Physcomitrella patens

    Directory of Open Access Journals (Sweden)

    Elizabeth A. Berry

    2016-03-01

    Full Text Available In contrast to homeohydric vascular plants, mosses employ a poikilohydric strategy for surviving in the dry aerial environment. A detailed understanding of the structure, composition, and development of moss cell walls can contribute to our understanding of not only the evolution of overall cell wall complexity, but also the differences that have evolved in response to selection for different survival strategies. The model moss species Physcomitrella patens has a predominantly haploid lifecycle consisting of protonemal filaments that regenerate from protoplasts and enlarge by tip growth, and leafy gametophores composed of cells that enlarge by diffuse growth and differentiate into several different types. Advantages for genetic studies include methods for efficient targeted gene modification and extensive genomic resources. Immuno and affinity cytochemical labeling were used to examine the distribution of polysaccharides and proteins in regenerated protoplasts, protonemal filaments, rhizoids, and sectioned gametophores of P. patens. The cell wall composition of regenerated protoplasts was also characterized by flow cytometry. Crystalline cellulose was abundant in the cell walls of regenerating protoplasts and protonemal cells that developed on media of high osmolarity, whereas homogalacturonan was detected in the walls of protonemal cells that developed on low osmolarity media and not in regenerating protoplasts. Mannan was the major hemicellulose detected in all tissues tested. Arabinogalactan proteins were detected in different cell types by different probes, consistent with structural heterogeneity. The results reveal developmental and cell type specific differences in cell wall composition and provide a basis for analyzing cell wall phenotypes in knockout mutants.

  3. Immuno and Affinity Cytochemical Analysis of Cell Wall Composition in the Moss Physcomitrella patens.

    Science.gov (United States)

    Berry, Elizabeth A; Tran, Mai L; Dimos, Christos S; Budziszek, Michael J; Scavuzzo-Duggan, Tess R; Roberts, Alison W

    2016-01-01

    In contrast to homeohydric vascular plants, mosses employ a poikilohydric strategy for surviving in the dry aerial environment. A detailed understanding of the structure, composition, and development of moss cell walls can contribute to our understanding of not only the evolution of overall cell wall complexity, but also the differences that have evolved in response to selection for different survival strategies. The model moss species Physcomitrella patens has a predominantly haploid lifecycle consisting of protonemal filaments that regenerate from protoplasts and enlarge by tip growth, and leafy gametophores composed of cells that enlarge by diffuse growth and differentiate into several different types. Advantages for genetic studies include methods for efficient targeted gene modification and extensive genomic resources. Immuno and affinity cytochemical labeling were used to examine the distribution of polysaccharides and proteins in regenerated protoplasts, protonemal filaments, rhizoids, and sectioned gametophores of P. patens. The cell wall composition of regenerated protoplasts was also characterized by flow cytometry. Crystalline cellulose was abundant in the cell walls of regenerating protoplasts and protonemal cells that developed on media of high osmolarity, whereas homogalactuonan was detected in the walls of protonemal cells that developed on low osmolarity media and not in regenerating protoplasts. Mannan was the major hemicellulose detected in all tissues tested. Arabinogalactan proteins were detected in different cell types by different probes, consistent with structural heterogneity. The results reveal developmental and cell type specific differences in cell wall composition and provide a basis for analyzing cell wall phenotypes in knockout mutants. PMID:27014284

  4. Immuno and Affinity Cytochemical Analysis of Cell Wall Composition in the Moss Physcomitrella patens.

    Science.gov (United States)

    Berry, Elizabeth A; Tran, Mai L; Dimos, Christos S; Budziszek, Michael J; Scavuzzo-Duggan, Tess R; Roberts, Alison W

    2016-01-01

    In contrast to homeohydric vascular plants, mosses employ a poikilohydric strategy for surviving in the dry aerial environment. A detailed understanding of the structure, composition, and development of moss cell walls can contribute to our understanding of not only the evolution of overall cell wall complexity, but also the differences that have evolved in response to selection for different survival strategies. The model moss species Physcomitrella patens has a predominantly haploid lifecycle consisting of protonemal filaments that regenerate from protoplasts and enlarge by tip growth, and leafy gametophores composed of cells that enlarge by diffuse growth and differentiate into several different types. Advantages for genetic studies include methods for efficient targeted gene modification and extensive genomic resources. Immuno and affinity cytochemical labeling were used to examine the distribution of polysaccharides and proteins in regenerated protoplasts, protonemal filaments, rhizoids, and sectioned gametophores of P. patens. The cell wall composition of regenerated protoplasts was also characterized by flow cytometry. Crystalline cellulose was abundant in the cell walls of regenerating protoplasts and protonemal cells that developed on media of high osmolarity, whereas homogalactuonan was detected in the walls of protonemal cells that developed on low osmolarity media and not in regenerating protoplasts. Mannan was the major hemicellulose detected in all tissues tested. Arabinogalactan proteins were detected in different cell types by different probes, consistent with structural heterogneity. The results reveal developmental and cell type specific differences in cell wall composition and provide a basis for analyzing cell wall phenotypes in knockout mutants.

  5. Altered catecholamine receptor affinity in rabbit aortic intimal hyperplasia

    International Nuclear Information System (INIS)

    Intimal thickening is a universal response to endothelial denudation and is also thought to be a precursor of atherosclerosis. The authors have demonstrated selective supersensitivity in arterial intimal hyperplasia to norepinephrine and they now report a possible mechanism for this. Binding studies in rabbit aorta with the selective alpha 1-adrenergic radioligand 125I-HEAT demonstrated that there was no change in receptor density (20 ± 4 fmole/10(6) cells) in intact vascular smooth muscle cells at either 5 or 14 days after denudation. However, competition studies showed a 2.6-fold increase in alpha 1-adrenergic receptor affinity for norepinephrine in intimal hyperplastic tissue (P less than 0.05). This increased affinity for norepinephrine was associated with a greater increase in 32P-labeled phosphatidylinositol (148% intimal thickening versus 76% control) and phosphatidic acid (151% intimal thickening versus 56% control) following norepinephrine stimulation of free floating rings of intimal hyperplastic aorta. These data suggest that the catecholamine supersensitivity in rabbit aortic intimal hyperplasia is receptor mediated and may be linked to the phosphatidylinositol cycle

  6. Altered catecholamine receptor affinity in rabbit aortic intimal hyperplasia

    Energy Technology Data Exchange (ETDEWEB)

    O' Malley, M.K.; Cotecchia, S.; Hagen, P.O. (Duke University Medical Center, Durham, NC (USA))

    1991-08-01

    Intimal thickening is a universal response to endothelial denudation and is also thought to be a precursor of atherosclerosis. The authors have demonstrated selective supersensitivity in arterial intimal hyperplasia to norepinephrine and they now report a possible mechanism for this. Binding studies in rabbit aorta with the selective alpha 1-adrenergic radioligand 125I-HEAT demonstrated that there was no change in receptor density (20 {plus minus} 4 fmole/10(6) cells) in intact vascular smooth muscle cells at either 5 or 14 days after denudation. However, competition studies showed a 2.6-fold increase in alpha 1-adrenergic receptor affinity for norepinephrine in intimal hyperplastic tissue (P less than 0.05). This increased affinity for norepinephrine was associated with a greater increase in 32P-labeled phosphatidylinositol (148% intimal thickening versus 76% control) and phosphatidic acid (151% intimal thickening versus 56% control) following norepinephrine stimulation of free floating rings of intimal hyperplastic aorta. These data suggest that the catecholamine supersensitivity in rabbit aortic intimal hyperplasia is receptor mediated and may be linked to the phosphatidylinositol cycle.

  7. Duals of Affine Grassmann Codes and Their Relatives

    DEFF Research Database (Denmark)

    Beelen, P.; Ghorpade, S. R.; Hoholdt, T.

    2012-01-01

    Affine Grassmann codes are a variant of generalized Reed-Muller codes and are closely related to Grassmann codes. These codes were introduced in a recent work by Beelen Here, we consider, more generally, affine Grassmann codes of a given level. We explicitly determine the dual of an affine Grassm...

  8. The Yeast PH Domain Proteins Slm1 and Slm2 Are Targets of Sphingolipid Signaling during the Response to Heat Stress▿ †

    OpenAIRE

    Daquinag, Alexes; Fadri, Maria; Jung, Sung Yun; Qin, Jun; Kunz, Jeannette

    2006-01-01

    The PH domain-containing proteins Slm1 and Slm2 were previously identified as effectors of the phosphatidylinositol-4,5-bisphosphate (PI4,5P2) and TORC2 signaling pathways. Here, we demonstrate that Slm1 and Slm2 are also targets of sphingolipid signaling during the heat shock response. We show that upon depletion of cellular sphingolipid levels, Slm1 function becomes essential for survival under heat stress. We further demonstrate that Slm proteins are regulated by a phosphorylation/dephosph...

  9. Generation of recombinant antibodies to rat GABAA receptor subunits by affinity selection on synthetic peptides.

    Directory of Open Access Journals (Sweden)

    Sujatha P Koduvayur

    Full Text Available The abundance and physiological importance of GABAA receptors in the central nervous system make this neurotransmitter receptor an attractive target for localizing diagnostic and therapeutic biomolecules. GABAA receptors are expressed within the retina and mediate synaptic signaling at multiple stages of the visual process. To generate monoclonal affinity reagents that can specifically recognize GABAA receptor subunits, we screened two bacteriophage M13 libraries, which displayed human scFvs, by affinity selection with synthetic peptides predicted to correspond to extracellular regions of the rat α1 and β2 GABAA subunits. We isolated three anti-β2 and one anti-α1 subunit specific scFvs. Fluorescence polarization measurements revealed all four scFvs to have low micromolar affinities with their cognate peptide targets. The scFvs were capable of detecting fully folded GABAA receptors heterologously expressed by Xenopus laevis oocytes, while preserving ligand-gated channel activity. Moreover, A10, the anti-α1 subunit-specific scFv, was capable of detecting native GABAA receptors in the mouse retina, as observed by immunofluorescence staining. In order to improve their apparent affinity via avidity, we dimerized the A10 scFv by fusing it to the Fc portion of the IgG. The resulting scFv-Fc construct had a Kd of ∼26 nM, which corresponds to an approximately 135-fold improvement in binding, and a lower detection limit in dot blots, compared to the monomeric scFv. These results strongly support the use of peptides as targets for generating affinity reagents to membrane proteins and encourage investigation of molecular conjugates that use scFvs as anchoring components to localize reagents of interest at GABAA receptors of retina and other neural tissues, for studies of receptor activation and subunit structure.

  10. The C2 domains of granuphilin are high-affinity sensors for plasma membrane lipids.

    Science.gov (United States)

    Lyakhova, Tatyana A; Knight, Jefferson D

    2014-09-01

    Membrane-targeting proteins are crucial components of many cell signaling pathways, including the secretion of insulin. Granuphilin, also known as synaptotagmin-like protein 4, functions in tethering secretory vesicles to the plasma membrane prior to exocytosis. Granuphilin docks to insulin secretory vesicles through interaction of its N-terminal domain with vesicular Rab proteins; however, the mechanisms of granuphilin plasma membrane targeting and release are less clear. Granuphilin contains two C2 domains, C2A and C2B, that interact with the plasma membrane lipid phosphatidylinositol-(4,5)-bisphosphate [PI(4,5)P2]. The goal of this study was to determine membrane-binding mechanisms, affinities, and kinetics of both granuphilin C2 domains using fluorescence spectroscopic techniques. Results indicate that both C2A and C2B bind anionic lipids in a Ca(2+)-independent manner. The C2A domain binds liposomes containing a physiological mixture of lipids including 2% PI(4,5)P2 or PI(3,4,5)P3 with high affinity (apparent K(d, PIPx) of 2-5 nM), and binds nonspecifically with moderate affinity to anionic liposomes lacking phosphatidylinositol phosphate (PIPx) lipids. The C2B domain binds with sub-micromolar affinity to liposomes containing PI(4,5)P2 but does not have a measurable affinity for background anionic lipids. Both domains can be competed away from their target lipids by the soluble PIPx analog inositol-(1,2,3,4,5,6)-hexakisphosphate (IP6), which is a positive regulator of insulin secretion. Potential roles of these interactions in the docking and release of granuphilin from the plasma membrane are discussed.

  11. Epidermal transformation leads to increased perlecan synthesis with heparin-binding-growth-factor affinity

    DEFF Research Database (Denmark)

    Tapanadechopone, P; Tumova, S; Jiang, X;

    2001-01-01

    . Despite this, the heparan sulphate of RT101- and JB6-derived perlecan bound fibroblast growth factor-1, -2, -4 and -7 and heparin-binding epidermal growth factor with similar affinity. Therefore abundant tumour-derived perlecan may support the angiogenic responses seen in vivo and be a key player...

  12. Measurement of the Isoscalar Monopole Response in the Neutron-Rich Nucleus 68Ni using the Active Target MAYA

    Science.gov (United States)

    Vandebrouck, M.; Gibelin, J.; Khan, E.; Achouri, N. L.; Baba, H.; Beaumel, D.; Blumenfeld, Y.; Caamaño, M.; Càceres, L.; Colò, G.; Delaunay, F.; Fernandez-Dominguez, B.; Garg, U.; Grinyer, G. F.; Harakeh, M. N.; Kalantar-Nayestanaki, N.; Keeley, N.; Mittig, W.; Pancin, J.; Raabe, R.; Roger, T.; Roussel-Chomaz, P.; Savajols, H.; Sorlin, O.; Stodel, C.; Suzuki, D.; Thomas, J. C.

    We report the measurement of the isoscalar monopole strength in the unstable nucleus 68Ni using inelastic alpha scattering at 50A MeV in inverse kinematics. This experiment has been performed at GANIL with LISE spectrometer using a dedicated detector: the active target MAYA. A part of the isoscalar giant monopole resonance (ISGMR) has been measured at 21.1 ± 1.9 MeV and indications for a soft monopole mode are provided for the first time at 12.9 ± 1.0 MeV. Distorted-wave born approximation (DWBA) with random-phase approximation (RPA) transition densities have been used to study angular distribution and indicate that the L = 0 multipolarity dominates the cross-section for the ISGMR, and significantly contributes to the soft mode.

  13. Targeting Anabolic Impairment in Response to Resistance Exercise in Older Adults with Mobility Impairments: Potential Mechanisms and Rehabilitation Approaches

    OpenAIRE

    Drummond, Micah J.; Robin L. Marcus; LaStayo, Paul C.

    2012-01-01

    Muscle atrophy is associated with healthy aging (i.e., sarcopenia) and may be compounded by comorbidities, injury, surgery, illness, and physical inactivity. While a bout of resistance exercise increases protein synthesis rates in healthy young skeletal muscle, the effectiveness of resistance exercise to mount a protein synthetic response is less pronounced in older adults. Improving anabolic sensitivity to resistance exercise, thereby enhancing physical function, is most critical in needy ol...

  14. Targeting Anabolic Impairment in Response to Resistance Exercise in Older Adults with Mobility Impairments: Potential Mechanisms and Rehabilitation Approaches

    Directory of Open Access Journals (Sweden)

    Micah J. Drummond

    2012-01-01

    Full Text Available Muscle atrophy is associated with healthy aging (i.e., sarcopenia and may be compounded by comorbidities, injury, surgery, illness, and physical inactivity. While a bout of resistance exercise increases protein synthesis rates in healthy young skeletal muscle, the effectiveness of resistance exercise to mount a protein synthetic response is less pronounced in older adults. Improving anabolic sensitivity to resistance exercise, thereby enhancing physical function, is most critical in needy older adults with clinical conditions that render them “low responders”. In this paper, we discuss potential mechanisms contributing to anabolic impairment to resistance exercise and highlight the need to improve anabolic responsiveness in low responders. This is followed with evidence suggesting that the recovery period of resistance exercise provides an opportunity to amplify the exercise-induced anabolic response using protein/essential amino acid ingestion. This anabolic strategy, if repeated chronically, may improve lean muscle gains, decrease time to recovery of function during periods of rehabilitation, and overall, maintain/improve physical independence and reduce mortality rates in older adults.

  15. Targeted Overexpression of Inducible 6-Phosphofructo-2-kinase in Adipose Tissue Increases Fat Deposition but Protects against Diet-induced Insulin Resistance and Inflammatory Responses*

    Science.gov (United States)

    Huo, Yuqing; Guo, Xin; Li, Honggui; Xu, Hang; Halim, Vera; Zhang, Weiyu; Wang, Huan; Fan, Yang-Yi; Ong, Kuok Teong; Woo, Shih-Lung; Chapkin, Robert S.; Mashek, Douglas G.; Chen, Yanming; Dong, Hui; Lu, Fuer; Wei, Lai; Wu, Chaodong

    2012-01-01

    Increasing evidence demonstrates the dissociation of fat deposition, the inflammatory response, and insulin resistance in the development of obesity-related metabolic diseases. As a regulatory enzyme of glycolysis, inducible 6-phosphofructo-2-kinase (iPFK2, encoded by PFKFB3) protects against diet-induced adipose tissue inflammatory response and systemic insulin resistance independently of adiposity. Using aP2-PFKFB3 transgenic (Tg) mice, we explored the ability of targeted adipocyte PFKFB3/iPFK2 overexpression to modulate diet-induced inflammatory responses and insulin resistance arising from fat deposition in both adipose and liver tissues. Compared with wild-type littermates (controls) on a high fat diet (HFD), Tg mice exhibited increased adiposity, decreased adipose inflammatory response, and improved insulin sensitivity. In a parallel pattern, HFD-fed Tg mice showed increased hepatic steatosis, decreased liver inflammatory response, and improved liver insulin sensitivity compared with controls. In both adipose and liver tissues, increased fat deposition was associated with lipid profile alterations characterized by an increase in palmitoleate. Additionally, plasma lipid profiles also displayed an increase in palmitoleate in HFD-Tg mice compared with controls. In cultured 3T3-L1 adipocytes, overexpression of PFKFB3/iPFK2 recapitulated metabolic and inflammatory changes observed in adipose tissue of Tg mice. Upon treatment with conditioned medium from iPFK2-overexpressing adipocytes, mouse primary hepatocytes displayed metabolic and inflammatory responses that were similar to those observed in livers of Tg mice. Together, these data demonstrate a unique role for PFKFB3/iPFK2 in adipocytes with regard to diet-induced inflammatory responses in both adipose and liver tissues. PMID:22556414

  16. Analysis of affinely equivalent Boolean functions

    Institute of Scientific and Technical Information of China (English)

    MENG QingShu; ZHANG HuanGuo; YANG Min; WANG ZhangYi

    2007-01-01

    By some basic transforms and invariant theory, we give two results: 1) an algorithm,which can be used to judge if two Boolean functions are affinely equivalent and to obtain the equivalence relationship if they are equivalent. This is useful in studying Boolean functions and in engineering. For example, we classify all 8-variable homogeneous bent functions of degree 3 into two classes; 2) Reed-Muller codes R(4,6)/R(1,6), R(3,7)/R(1,7) are classified efficiently.

  17. On Metrizability of Invariant Affine Connections

    CERN Document Server

    Tanaka, Erico

    2011-01-01

    The metrizability problem for a symmetric affine connection on a manifold, invariant with respect to a group of diffeomorphisms G, is considered. We say that the connection is G-metrizable, if it is expressible as the Levi-Civita connection of a G-invariant metric field. In this paper we analyze the G-metrizability equations for the rotation group G = SO(3), acting canonically on three- and four-dimensional Euclidean spaces. We show that the property of the connection to be SO(3)-invariant allows us to find complete explicit description of all solutions of the SO(3)-metrizability equations.

  18. Latest European coelacanth shows Gondwanan affinities.

    Science.gov (United States)

    Cavin, Lionel; Forey, Peter L; Buffetaut, Eric; Tong, Haiyan

    2005-06-22

    The last European fossil occurrence of a coelacanth is from the Mid-Cretaceous of the English Chalk (Turonian, 90 million years ago). Here, we report the discovery of a coelacanth from Late Cretaceous non-marine rocks in southern France. It consists of a left angular bone showing structures that imply close phylogenetic affinities with some extinct Mawsoniidae. The closest relatives are otherwise known from Cretaceous continental deposits of southern continents and suggest that the dispersal of freshwater organisms from Africa to Europe occurred in the Late Cretaceous.

  19. Affine Coherent States in Quantum Cosmology

    CERN Document Server

    Malkiewicz, Przemyslaw

    2015-01-01

    A brief summary of the application of coherent states in the examination of quantum dynamics of cosmological models is given. We discuss quantization maps, phase space probability distributions and semiclassical phase spaces. The implementation of coherent states based on the affine group resolves the hardest singularities, renders self-adjoint Hamiltonians without boundary conditions and provides a completely consistent semi-classical description of the involved quantum dynamics. We consider three examples: the closed Friedmann model, the anisotropic Bianchi Type I model and the deep quantum domain of the Bianchi Type IX model.

  20. Measuring an antibody affinity distribution molecule by molecule

    Energy Technology Data Exchange (ETDEWEB)

    Bradbury, Andrew M [Los Alamos National Laboratory; Werner, James H [Los Alamos National Laboratory; Temirov, Jamshid [INVITROGEN

    2008-01-01

    Single molecule fluorescence mIcroscopy was used to observe the binding and unbinding of hapten decorated quantum dots with individual surface immobilized antibodies. The fluorescence time history from an individual antibody site can be used to calculate its binding affinity. While quantum dot blinking occurs during these measurements, we describe a simple empirical method to correct the apparent/observed affinity to account for the blinking contribution. The combination of many single molecule affinity measurements from different antibodies yields not only the average affinity, it directly measures the full shape and character of the surface affinity distribution function.