Submillimeter Follow-up of WISE-Selected Hyperluminous Galaxies

We have used the Caltech Submillimeter Observatory (CSO) to follow-up a sample of WISE-selected, hyperluminous galaxies, so called W1W2-dropout galaxies. This is a rare (~ 1000 all-sky) population of galaxies at high redshift (peaks at z=2-3), that are faint or undetected by WISE at 3.4 and 4.6 um, yet are clearly detected at 12 and 22 um. The optical spectra of most of these galaxies show significant AGN activity. We observed 14 high-redshift (z > 1.7) W1W2-dropout galaxies with SHARC-II at 350 to 850 um, with 9 detections; and observed 18 with Bolocam at 1.1 mm, with five detections. Warm Spitzer follow-up of 25 targets at 3.6 and 4.5 um, as well as optical spectra of 12 targets are also presented in the paper. Combining WISE data with observations from warm Spitzer and CSO, we constructed their mid-IR to millimeter spectral energy distributions (SEDs). These SEDs have a consistent shape, showing significantly higher mid-IR to submm ratios than other galaxy templates, suggesting a hotter dust temperature. We estimate their dust temperatures to be 60-120 K using a single-temperature model. Their infrared luminosities are well over 10^{13} Lsun. These SEDs are not well fitted with existing galaxy templates, suggesting they are a new population with very high luminosity and hot dust. They are likely among the most luminous galaxies in the Universe. We argue that they are extreme cases of luminous, hot dust-obscured galaxies (DOGs), possibly representing a short evolutionary phase during galaxy merging and evolution. A better understanding of their long-wavelength properties needs ALMA as well as Herschel data.Comment: Will be Published on Sep 1, 2012 by Ap

Cyanine dye mediated mitochondrial targeting enhances the anti-cancer activity of small-molecule cargoes

Organelle-specific delivery systems are of significant clinical interest. We demonstrate the use of common cyanine dyes Cy3 and Cy5 as vectors for targeting and delivering cargoes to mitochondria in cancer cells. Specifically, conjugation to the dyes can increase cytotoxicity by up to 1000-fold

Stereoisomer-dependent membrane association and capacity for insulin delivery facilitated by Penetratin

Cell-penetrating peptides (CPPs), such as penetratin, are often investigated as drug delivery vectors and incorporating d-amino acids, rather than the natural l-forms, to enhance proteolytic stability could improve their delivery efficiency. The present study aimed to compare membrane association, cellular uptake, and delivery capacity for all-l and all-d enantiomers of penetratin (PEN) by using different cell models and cargos. The enantiomers displayed widely different distribution patterns in the examined cell models, and in Caco-2 cells, quenchable membrane binding was evident for d-PEN in addition to vesicular intracellular localization for both enantiomers. The uptake of insulin in Caco-2 cells was equally mediated by the two enantiomers, and while l-PEN did not increase the transepithelial permeation of any of the investigated cargo peptides, d-PEN increased the transepithelial delivery of vancomycin five-fold and approximately four-fold for insulin at an extracellular apical pH of 6.5. Overall, while d-PEN was associated with the plasma membrane to a larger extent and was superior in mediating the transepithelial delivery of hydrophilic peptide cargoes compared to l-PEN across Caco-2 epithelium, no enhanced delivery of the hydrophobic cyclosporin was observed, and intracellular insulin uptake was induced to a similar degree by the two enantiomers

Targeted delivery and endosomal cellular uptake of DARPin-siRNA bioconjugates: Influence of linker stability on gene silencing

Specific cell targeting and efficient intracellular delivery are major hurdles for the widespread therapeutic use of nucleic acid technologies, particularly siRNA mediated gene silencing. To enable receptor-mediated cell-specific targeting, we designed a synthesis scheme that can be generically used to engineer Designed Ankyrin Repeat Protein (DARPin)-siRNA bioconjugates. Different linkers, including labile disulfide-, and more stable thiol-maleimide- and triazole- (click chemistry) tethers were employed. Crosslinkers were first attached to a 3’-terminal aminohexyl chain on the siRNA sense strands. On the protein side thiols of a C-terminal cysteine were used as anchoring site for disulfide- and thiol-maleimide conjugate formation, while strain-promoted azido-alkyne cycloadditions were carried out at a metabolically introduced N-terminal azidohomoalanine. After establishing efficient purification methods, highly pure products were obtained. Bioconjugates of EpCAM-targeted DARPins with siRNA directed at the luciferase gene were evaluated for cell-specific binding, uptake and gene silencing. As shown by flow cytometry and fluorescence microscopy, all constructs retained the highly specific and high-affinity antigen recognition properties of the native DARPin. As expected, internalization was observed only in EpCAM-positive cell lines, and predominantly endolysosomal localization was detected. Disulfide linked conjugates showed lower serum stability against cleavage at the linker and thus lower internalization into endosomes compared to thiol-maleimide- and triazole-linked conjugates, yet induced more pronounced gene silencing. This indicates that the siRNA payload needs to be liberated from the protein in the endosome. Our data confirm the promise of DARPin-siRNA bioconjugates for tumor targeting, but also identified endosomal retention and limited cytosolic escape of the siRNA as the rate-limiting step for more efficient gene silencing

Four-wave-mixing microscopy reveals non-colocalisation between gold nanoparticles and fluorophore conjugates inside cells

Gold nanoparticles have been researched for many biomedical applications in diagnostics, theranostics, and as drug delivery systems. When conjugated to fluorophores, their interaction with biological cells can be studied in situ and real time using fluorescence microscopy. However, an important question that has remained elusive to answer is whether the fluorophore is a faithful reporter of the nanoparticle location. Here, our recently developed four-wave-mixing optical microscopy is applied to image individual gold nanoparticles and in turn investigate their co-localisation with fluorophores inside cells. Nanoparticles from 10 nm to 40 nm diameter were conjugated to fluorescently-labeled transferrin, for internalisation via clathrin-mediated endocytosis, or to non-targeting fluorescently-labelled antibodies. Human (HeLa) and murine (3T3-L1) cells were imaged at different time points after incubation with these conjugates. Our technique identified that, in most cases, fluorescence originated from unbound fluorophores rather than from fluorophores attached to nanoparticles. Fluorescence detection was also severely limited by photobleaching, quenching and autofluorescence background. Notably, correlative extinction/fluorescence microscopy of individual particles on a glass surface indicated that commercial constructs contain large amounts of unbound fluorophores. These findings highlight the potential problems of data interpretation when reliance is solely placed on the detection of fluorescence within the cell, and are of significant importance in the context of correlative light electron microscopy

Highly luminescent metallacages featuring bispyridyl ligands functionalised with BODIPY for imaging in cancer cells

Recently, 3-dimentional supramolecular coordination complexes of the metallacage type have been shown to hold promise as drug delivery systems for different cytotoxic agents, including the anticancer drug cisplatin. However, so far only limited information is available on their uptake and sub-cellular localisation in cancer cells. With the aim of understanding the fate of metallacages in cells by fluorescence microscopy, three fluorescent Pd2L4 metallacages were designed and synthesised by self-assembly of two types of bispyridyl ligands (L), exo-functionalised with boron dipyrromethene (BODIPY) moieties, with Pd(II) ions. The cages show high quantum yields and are moderately stable in the presence of physiologically relevant concentration of glutathione. Furthermore, the cages are able to encapsulate the anticancer drug cisplatin, as demonstrated by NMR spectroscopy. Preliminary cytotoxicity studies in a small panel of human cancer cells showed that the metallacages are scarcely toxic in vitro. The marked fluorescence due to BODIPY allowed us to visualise the cages' uptake and sub-cellular localisation inside melanoma cells using fluorescence microscopy, highlighting uptake via active transport mechanisms and accumulation in cytoplasmic vesicles

Exome-wide analysis of copy number variation shows association of the human leukocyte antigen region with asthma in UK Biobank

BackgroundThe role of copy number variants (CNVs) in susceptibility to asthma is not well understood. This is, in part, due to the difficulty of accurately measuring CNVs in large enough sample sizes to detect associations. The recent availability of whole-exome sequencing (WES) in large biobank studies provides an unprecedented opportunity to study the role of CNVs in asthma.MethodsWe called common CNVs in 49,953 individuals in the first release of UK Biobank WES using ClinCNV software. CNVs were tested for association with asthma in a stage 1 analysis comprising 7098 asthma cases and 36,578 controls from the first release of sequencing data. Nominally-associated CNVs were then meta-analysed in stage 2 with an additional 17,280 asthma cases and 115,562 controls from the second release of UK Biobank exome sequencing, followed by validation and fine-mapping.ResultsFive of 189 CNVs were associated with asthma in stage 2, including a deletion overlapping the HLA-DQA1 and HLA-DQB1 genes, a duplication of CHROMR/PRKRA, deletions within MUC22 and TAP2, and a duplication in FBRSL1. The HLA-DQA1, HLA-DQB1, MUC22 and TAP2 genes all reside within the human leukocyte antigen (HLA) region on chromosome 6. In silico analyses demonstrated that the deletion overlapping HLA-DQA1 and HLA-DQB1 is likely to be an artefact arising from under-mapping of reads from non-reference HLA haplotypes, and that the CHROMR/PRKRA and FBRSL1 duplications represent presence/absence of pseudogenes within the HLA region. Bayesian fine-mapping of the HLA region suggested that there are two independent asthma association signals. The variants with the largest posterior inclusion probability in the two credible sets were an amino acid change in HLA-DQB1 (glutamine to histidine at residue 253) and a multi-allelic amino acid change in HLA-DRB1 (presence/absence of serine, glycine or leucine at residue 11).ConclusionsAt least two independent loci characterised by amino acid changes in the HLA-DQA1, HLA-DQB1 and HLA-DRB1 genes are likely to account for association of SNPs and CNVs in this region with asthma. The high divergence of haplotypes in the HLA can give rise to spurious CNVs, providing an important, cautionary tale for future large-scale analyses of sequencing data

Copy number variation of the beta-defensin genes in Europeans: no supporting evidence for association with lung function, chronic obstructive pulmonary disease or asthma

Lung function measures are heritable, predict mortality and are relevant in diagnosis of chronic obstructive pulmonary disease (COPD). COPD and asthma are diseases of the airways with major public health impacts and each have a heritable component. Genome-wide association studies of SNPs have revealed novel genetic associations with both diseases but only account for a small proportion of the heritability. Complex copy number variation may account for some of the missing heritability. A well-characterised genomic region of complex copy number variation contains beta-defensin genes (DEFB103, DEFB104 and DEFB4), which have a role in the innate immune response. Previous studies have implicated these and related genes as being associated with asthma or COPD. We hypothesised that copy number variation of these genes may play a role in lung function in the general population and in COPD and asthma risk. We undertook copy number typing of this locus in 1149 adult and 689 children using a paralogue ratio test and investigated association with COPD, asthma and lung function. Replication of findings was assessed in a larger independent sample of COPD cases and smoking controls. We found evidence for an association of beta-defensin copy number with COPD in the adult cohort (OR = 1.4, 95%CI:1.02–1.92, P = 0.039) but this finding, and findings from a previous study, were not replicated in a larger follow-up sample(OR = 0.89, 95%CI:0.72–1.07, P = 0.217). No robust evidence of association with asthma in children was observed. We found no evidence for association between beta-defensin copy number and lung function in the general populations. Our findings suggest that previous reports of association of beta-defensin copy number with COPD should be viewed with caution. Suboptimal measurement of copy number can lead to spurious associations. Further beta-defensin copy number measurement in larger sample sizes of COPD cases and children with asthma are needed

The First Hyper-luminous Infrared Galaxy Discovered by WISE

We report the discovery by the Wide-field Infrared Survey Explorer (WISE) of the z = 2.452 source WISE J181417.29+341224.9, the first hyperluminous source found in the WISE survey. WISE 1814+3412 is also the prototype for an all-sky sample of ~1000 extremely luminous "W1W2-dropouts" (sources faint or undetected by WISE at 3.4 and 4.6 μm and well detected at 12 or 22 μm). The WISE data and a 350 μm detection give a minimum bolometric luminosity of 3.7 × 10^(13) L_☉, with ~10^(14) L_☉ plausible. Follow-up images reveal four nearby sources: a QSO and two Lyman break galaxies (LBGs) at z = 2.45, and an M dwarf star. The brighter LBG dominates the bolometric emission. Gravitational lensing is unlikely given the source locations and their different spectra and colors. The dominant LBG spectrum indicates a star formation rate ~300 M_☉ yr^(–1), accounting for ≲ 10% of the bolometric luminosity. Strong 22 μm emission relative to 350 μm implies that warm dust contributes significantly to the luminosity, while cooler dust normally associated with starbursts is constrained by an upper limit at 1.1 mm. Radio emission is ~10 times above the far-infrared/radio correlation, indicating an active galactic nucleus (AGN) is present. An obscured AGN combined with starburst and evolved stellar components can account for the observations. If the black hole mass follows the local M BH-bulge mass relation, the implied Eddington ratio is ≳ 4. WISE 1814+3412 may be a heavily obscured object where the peak AGN activity occurred prior to the peak era of star formation