CXCR4-targeted and MMP-responsive iron oxide nanoparticles for enhanced magnetic resonance imaging

MRI offers high spatial resolution with excellent tissue penetration but it has limited sensitivity and the commonly administered contrast agents lack specificity. In this study, two sets of iron oxide nanoparticles (IONPs) were synthesized that were designed to selectively undergo copper-free click conjugation upon sensing of matrix metalloproteinase (MMP) enzymes, thereby leading to a self-assembled superparamagnetic nanocluster network with T2 signal enhancement properties. For this purpose, IONPs with bioorthogonal azide and alkyne surfaces masked by polyethylene glycol (PEG) layers tethered to CXCR4-targeted peptide ligands were synthesized and characterized. The IONPs were tested in vitro and T2 signal enhancements of around 160 % were measured when the IONPs were incubated with cells expressing MMP2/9 and CXCR4. Simultaneous systemic administration of the bioorthogonal IONPs in tumor-bearing mice demonstrated the signal-enhancing ability of these ‘smart’ self-assembling nanomaterials

Multi-Frame Rate Plane Wave Contrast-Enhance Ultrasound Imaging for Tumour Vasculature Imaging and Perfusion Quantification

A multi-frame rate plane wave imaging strategy is developed to simultaneously image tumor vasculature and quantify tumor perfusion. Customised imaging sequences interleaving a short but high frame rate (HFR) plane wave imaging sequence with a long but low frame rate imaging (LFR) sequence were implemented using a programmable ultrasound research platform. The results from a spatio-temporal coherence processing technique of ours demonstrated a significant improvement in the SNR and vasculature contrast when compared with the existing ultrafast Power Doppler (PD) using the same data. Initial perfusion quantification using LFR imaging was also demonstrated. Mean time intensity curve and some parametric measures were generated. Combining both structural and functional perfusion imaging using the multiframe rate sequences, a better evaluation of the tumour angiogenesis can be assessed

Multi-frame rate plane wave contrast-enhanced ultrasound imaging for tumour vascular imaging and perfusion quantification

Angiogenesis and blood flow dynamics play an important role in the development of malignant tumours and their response to treatment. While contrast enhanced ultrasound (CEUS) imaging with microbubble contrast agents as a tool for imaging angiogenesis and flow dynamics has shown great potential [1], recent development of plane wave high frame-rate (HFR) CEUS has offered new opportunities in such applications. In this study, we demonstrate an interleaved multi-frame rate plane wave CEUS imaging to quantify perfusion and to image vascular structure with improved resolution and contrast

Investigating CXCR4 expression of tumor cells and the vascular compartment: A multimodal approach

The C-X-C chemokine receptor 4 (CXCR4) is G protein-coupled receptor that upon binding to its cognate ligand, can lead to tumor progression. Several CXCR4-targeted therapies are currently under investigation, and with it comes the need for imaging agents capable of accurate depiction of CXCR4 for therapeutic stratification and monitoring. PET agents enjoy the most success, but more cost-effective and radiation-free approaches such as ultrasound (US) imaging could represent an attractive alternative. In this work, we developed a targeted microbubble (MB) for imaging of vascular CXCR4 expression in cancer. A CXCR4-targeted MB was developed through incorporation of the T140 peptide into the MB shell. Binding properties of the T140-MB and control, non-targeted MB (NT-MB) were evaluated in MDA-MB-231 cells where CXCR4 expression was knocked-down (via shRNA) through optical imaging, and in the lymphoma tumor models U2932 and SuDHL8 (high and low CXCR4 expression, respectively) by US imaging. PET imaging of [18F]MCFB, a tumor-penetrating CXCR4-targeted small molecule, was used to provide whole-tumor CXCR4 readouts. CXCR4 expression and microvessel density were performed by immunohistochemistry analysis and western blot. T140-MB were formed with similar properties to NT-MB and accumulated sensitively and specifically in cells according to their CXCR4 expression. In NOD SCID mice, T140-MB persisted longer in tumors than NT-MB, indicative of target interaction, but showed no difference between U2932 and SuDHL8. In contrast, PET imaging with [18F]MCFB showed a marked difference in tumor uptake at 40–60 min post-injection between the two tumor models (p<0.05). Ex vivo analysis revealed that the large differences in CXCR4 expression between the two models are not reflected in the vascular compartment, where the MB are restricted; in fact, microvessel density and CXCR4 expression in the vasculature was comparable between U2932 and SuDHL8 tumors. In conclusion, we successfully developed a T140-MB that can be used for imaging CXCR4 expression in the tumor vasculature

The Turkey Ig-like receptor family: identification, expression and function.

The chicken leukocyte receptor complex located on microchromosome 31 encodes the chicken Ig-like receptors (CHIR), a vastly expanded gene family which can be further divided into three subgroups: activating CHIR-A, bifunctional CHIR-AB and inhibitory CHIR-B. Here, we investigated the presence of CHIR homologues in other bird species. The available genome databases of turkey, duck and zebra finch were screened with different strategies including BLAST searches employing various CHIR sequences, and keyword searches. We could not identify CHIR homologues in the distantly related zebra finch and duck, however, several partial and complete sequences of CHIR homologues were identified on chromosome 3 of the turkey genome. They were designated as turkey Ig-like receptors (TILR). Using cDNA derived from turkey blood and spleen RNA, six full length TILR could be amplified and further divided according to the typical sequence features into one activating TILR-A, one inhibitory TILR-B and four bifunctional TILR-AB. Since the TILR-AB sequences all displayed the critical residues shown to be involved in binding to IgY, we next confirmed the IgY binding using a soluble TILR-AB1-huIg fusion protein. This fusion protein reacted with IgY derived from various gallinaceous birds, but not with IgY from other bird species. Finally, we tested various mab directed against CHIR for their crossreactivity with either turkey or duck leukocytes. Whereas no staining was detectable with duck cells, the CHIR-AB1 specific mab 8D12 and the CHIR-A2 specific mab 13E2 both reacted with a leukocyte subpopulation that was further identified as thrombocytes by double immunofluorescence employing B-cell, T-cell and thrombocyte specific reagents. In summary, although the turkey harbors similar LRC genes as the chicken, their distribution seems to be distinct with predominance on thrombocytes rather than lymphocytes

Matched sizes of activating and inhibitory receptor/ligand pairs are required for optimal signal integration by human Natural Killer cells

It has been suggested that receptor-ligand complexes segregate or co-localise within immune synapses according to their size, and this is important for receptor signaling. Here, we set out to test the importance of receptor-ligand complex dimensions for immune surveillance of target cells by human Natural Killer (NK) cells. NK cell activation is regulated by integrating signals from activating receptors, such as NKG2D, and inhibitory receptors, such as KIR2DL1. Elongating the NKG2D ligand MICA reduced its ability to trigger NK cell activation. Conversely, elongation of KIR2DL1 ligand HLA-C reduced its ability to inhibit NK cells. Whereas normal-sized HLA-C was most effective at inhibiting activation by normal-length MICA, only elongated HLA-C could inhibit activation by elongated MICA. Moreover, HLA-C and MICA that were matched in size co-localised, whereas HLA-C and MICA that were different in size were segregated. These results demonstrate that receptor-ligand dimensions are important in NK cell recognition, and suggest that optimal integration of activating and inhibitory receptor signals requires the receptor-ligand complexes to have similar dimensions

Modulation of enhancer looping and differential gene targeting by Epstein-Barr virus transcription factors directs cellular reprogramming

Epstein-Barr virus (EBV) epigenetically reprogrammes B-lymphocytes to drive immortalization and facilitate viral persistence. Host-cell transcription is perturbed principally through the actions of EBV EBNA 2, 3A, 3B and 3C, with cellular genes deregulated by specific combinations of these EBNAs through unknown mechanisms. Comparing human genome binding by these viral transcription factors, we discovered that 25% of binding sites were shared by EBNA 2 and the EBNA 3s and were located predominantly in enhancers. Moreover, 80% of potential EBNA 3A, 3B or 3C target genes were also targeted by EBNA 2, implicating extensive interplay between EBNA 2 and 3 proteins in cellular reprogramming. Investigating shared enhancer sites neighbouring two new targets (WEE1 and CTBP2) we discovered that EBNA 3 proteins repress transcription by modulating enhancer-promoter loop formation to establish repressive chromatin hubs or prevent assembly of active hubs. Re-ChIP analysis revealed that EBNA 2 and 3 proteins do not bind simultaneously at shared sites but compete for binding thereby modulating enhancer-promoter interactions. At an EBNA 3-only intergenic enhancer site between ADAM28 and ADAMDEC1 EBNA 3C was also able to independently direct epigenetic repression of both genes through enhancer-promoter looping. Significantly, studying shared or unique EBNA 3 binding sites at WEE1, CTBP2, ITGAL (LFA-1 alpha chain), BCL2L11 (Bim) and the ADAMs, we also discovered that different sets of EBNA 3 proteins bind regulatory elements in a gene and cell-type specific manner. Binding profiles correlated with the effects of individual EBNA 3 proteins on the expression of these genes, providing a molecular basis for the targeting of different sets of cellular genes by the EBNA 3s. Our results therefore highlight the influence of the genomic and cellular context in determining the specificity of gene deregulation by EBV and provide a paradigm for host-cell reprogramming through modulation of enhancer-promoter interactions by viral transcription factors

Support for a synaptic chain model of neuronal sequence generation

In songbirds, the remarkable temporal precision of song is generated by a sparse sequence of bursts in the premotor nucleus HVC. To distinguish between two possible classes of models of neural sequence generation, we carried out intracellular recordings of HVC neurons in singing zebra finches (Taeniopygia guttata). We found that the subthreshold membrane potential is characterized by a large, rapid depolarization 5–10 ms before burst onset, consistent with a synaptically connected chain of neurons in HVC. We found no evidence for the slow membrane potential modulation predicted by models in which burst timing is controlled by subthreshold dynamics. Furthermore, bursts ride on an underlying depolarization of ~10-ms duration, probably the result of a regenerative calcium spike within HVC neurons that could facilitate the propagation of activity through a chain network with high temporal precision. Our results provide insight into the fundamental mechanisms by which neural circuits can generate complex sequential behaviours.National Institutes of Health (U.S.) (Grant MH067105)National Institutes of Health (U.S.) (Grant DC009280)National Science Foundation (U.S.) (IOS-0827731)Alfred P. Sloan Foundation (Research Fellowship

The Effects of Biogeography on Ant Diversity and Activity on the Boston Harbor Islands, Massachusetts, U.S.A

Many studies have examined how island biogeography affects diversity on the scale of island systems. In this study, we address how diversity varies over very short periods of time on individual islands. To do this, we compile an inventory of the ants living in the Boston Harbor Islands National Recreation Area, Boston, Massachusetts, USA using data from a five-year All Taxa Biodiversity Inventory of the region's arthropods. Consistent with the classical theory of island biogeography, species richness increased with island size, decreased with island isolation, and remained relatively constant over time. Additionally, our inventory finds that almost half of the known Massachusetts ant fauna can be collected in the BHI, and identifies four new species records for Massachusetts, including one new to the United States, Myrmica scabrinodis. We find that the number of species actually active on islands depended greatly on the timescale under consideration. The species that could be detected during any given week of sampling could by no means account for total island species richness, even when correcting for sampling effort. Though we consistently collected the same number of species over any given week of sampling, the identities of those species varied greatly between weeks. This variation does not result from local immigration and extinction of species, nor from seasonally-driven changes in the abundance of individual species, but rather from weekly changes in the distribution and activity of foraging ants. This variation can be upwards of 50% of ant species per week. This suggests that numerous ant species on the BHI share the same physical space at different times. This temporal partitioning could well explain such unexpectedly high ant diversity in an isolated, urban site