Technical Reports in 2012

The glomerulus, unlike many capillary beds, readily supports leukocyte recruitment, but little is known about the actions of leukocytes upon recruitment to glomeruli. Devi and colleagues have addressed this issue with a multiphoton, confocal microscopy–based approach to shed light on glomerular inflammation. Induction of inflammation in mice did not increase the number of leukocytes recruited to glomeruli but prolonged the duration of leukocyte retention and migration.

Currently there is no single imaging system that can offer adequate spatial and temporal resolution to accurately assess many of the important factors involved in peripheral arterial diseases. Here, an epifluorescence imaging approach is offered that overcomes many of the current limitations and uses the near-infrared fluorescence of single-walled carbon nanotubes as fluorophores in the second near-infrared window (beyond 1,000 nm). Its use is demonstrated for imaging blood vessels in mouse hindlimb vasculatures millimeters deep in vivo.

Estrogen is beneficial for obesity and type 2 diabetes, though its use is limited by important side effects. In a new study, Matthias Tschöp and colleagues avoid this issue by chemically linking estrogen to the hormone GLP-1 to selectively target metabolically relevant tissue and show that the conjugated compound corrects obesity, hyperglycemia and dyslipidemia in mice. This approach could be used for other hormone pairs to treat other diseases.

Cancer cells shed large numbers of small, membrane-bound microvesicles (MVs) into the circulation, which have diagnostic potential but have proved difficult to analyze in a point-of-care setting. Huilin Shao and colleagues have developed a microfluidic chip with an integrated NMR detection system for the rapid profiling of circulating MVs directly from blood samples of patients with glioblastoma. The system was used to distinguish cancer cell–derived MVs from host cell–derived MVs and to measure treatment effects in vivo.

Chronic obstructive pulmonary disease (COPD) is now known to be a heterogeneous disorder, hence the pressing need to develop imaging biomarkers to differentiate between the broad range of COPD phenotypes–all of which require different treatments. Here, Craig Galban and his colleagues have adapted the parametric response map technique for the voxel-by-voxel classification analysis of CT lung images taken from a national COPD trial, providing a more objective characterization of the phenotypic contributions of functional small airways disease and emphysema in COPD.

By exploiting the relationship between the transcription factor MYC and the transferrin receptor, where the level of transferrin receptor 1 expression may indicate activation of the MYC oncogenic pathway, Jason Holland and his colleagues have developed a novel PET radiotracer to quantitatively and noninvasively measure MYC activity. The ⁸⁹Zr-desferrioxamine transferrin PET radiotracer was tested in several murine models of inflammation and MYC-driven prostate cancer.

A limitation of photodynamic therapy (PDT) is the depth of penetration of visible light needed for activation of the photosensitizers, restricting treatment to tumors on or just under the skin’s surface or those lining internal organs and cavities. Niagara Muhammad Idris and colleagues have addressed this issue by developing upconversion fluorescent nanoparticles (UNCs) that convert deeper penetrating near-infrared light to visible wavelengths without sacrificing efficacy for singlet oxygen (1O2) production. The group tested the UNCs in vivo in a subcutaneous mouse tumor model using a dual-sensitizer approach for greater PDT efficacy.

Using the semiconductor synthesis technology of maskless photolithography on microprocessor-grade silicon wafers, Jordan Price and colleagues synthesized microarrays containing every possible overlapping peptide in a linear sequence covering the N terminus of human histone H2B, including post-translational modifications. They demonstrated use of the 'on silico' peptide microarrays for the high-resolution mapping (at the single amino acid level) of epitopes targeted by commercially available H2B-specific antibodies and also by autoantibodies in samples from individuals with systemic lupus erythematosus.

Joon-Mo Yang and colleagues have developed a new endoscopic technique for the in vivo imaging of internal organs, combining endoscopic ultrasound and photoacoustic endoscopy in a single instrument. In addition to improved resolution, imaging depth, multimodal contrast, and distal-end scanning, the new hybrid imaging modality can also provide functional information such as hemoglobin concentration and blood oxygenation. Feasibility is shown in vivo by simultaneous photoacoustic endoscopy and endoscopic ultrasound imaging of the upper and lower gastrointestinal tracts of rats and rabbits.

The mucosa of the large intestine mucosa is an effective vaccination site for induction of protective mucosal immunity against rectal or vaginal viral infection but clinically is impractical. Here Qing Zhu et al. have developed an oral delivery system that encapsulates vaccine into pH-dependent poly(DL-lactic-co-glycolic acid) (PLGA) nanoparticles coated with Eudragit to protect against the low pH and enzymatic destruction of the stomach. This approach was shown to selectively target the large intestine in mice and induce antigen-specific T and B cell responses similar to that observed with intracolorectal vaccination.

In the past, small-diameter, synthetic vascular prostheses have been made in vitro either entirely from cells or by inoculating cells onto scaffolds. Wei Wu and colleagues have taken a cell-free approach where biodegradable elastomeric grafts are rapidly degraded, producing neoarteries that were almost free of foreign materials 90 days after grafting in a rat abdominal aorta. Grafts were rapidly remodeled by the host and produced compliance and burst pressure values similar to those of native aorta.

Nadia Dominici and her colleagues have developed a multidirectional robotic neurorehabilitation system that is capable of operating as a propulsive or postural neuroprosthesis and overcomes some of the limitations of existing systems. The robotic interface allows for the independent assessment and restoration of motor function in rats with mild to severe neuromotor disorders and is validated in various models of spinal cord injury and stroke.

For years, manufacturers have used one of only two chemicals to inactivate viruses for vaccine production: formaldehyde or β-propiolactone, and formaldehyde can damage key antigenic epitopes, leading to reduced immunogenicity or exacerbated disease. Ian Amanna and his colleagues have now found a third, the oxidizing agent hydrogen peroxide (H₂O₂), which they show can be more effective than the conventional approaches. Utility of the H₂O₂-based approach is demonstrated in three model systems.

T cell receptor (TCR)-based immunotherapeutic approaches have so far had limited success because of a lack of specific immune recognition and activation by the TCR. Here Nathaniel Liddy and his colleagues describe the generation, optimization and characterization of a new set of reagents—immune-mobilizing monoclonal TCRs against cancer (or ImmTACs)—designed to overcome some of these limitations. The ImmTACs were used to redirect and activate T cells to lyse tumor cells both in vitro and in vivo, even those expressing very low epitope numbers on the cell surface.

The ability to effectively assess tumor margins for brain tumor resection is a crucial factor in determining outcome in patients with brain tumors. Moritz Kircher and colleagues have developed a gold-silica nanoparticle that provides a triple-mode imaging capability of magnetic resonance, photoacoustic and Raman imaging, capitalizing on the complementary strengths of each modality for noninvasively delineating brain tumor margins both preoperatively and intraoperatively. The approach was tested in several mouse models, including one that recapitulates the infiltrating growth pattern of human gliomas.

Finding new methods to define the target antigens recognized by MHC class I–restricted T cells is an unmet need. Katherina Siewert and her colleagues have developed a sensitive technique based on recombinatorial plasmid screening of T cell receptors (TCRs) isolated from individual T cells that overcomes many of the current limitations and enables the characterization of T cell antigens from most T cells, including those isolated from frozen biopsy samples by laser microdissection. The approach was validated using a well-characterized influenza virus–specific TCR, MHC and peptide combination.

Building on their recent work establishing long-term cultures from Lgr5⁺ cells of the small intestine and stomach, Shiro Yui and colleagues describe a new colonic stem cell culture and expansion method generating organoids from single Lgr5⁺ stem cells. The study provides proof of principle that the cultured Lgr5⁺ cells can be used for stem cell therapy to repair superficially damaged epithelium in a dextran sulfate sodium (DSS)-induced acute colitis mouse model.

There are currently no good ways to track human cells in vivo in a clinical setting using magnetic resonance imaging (MRI) based on superparamagnetic iron-oxide nanoparticles. Mya Thu and colleagues have introduced a simple magnetic cell labeling approach that combines three currently US Food and Drug Administration–approved drugs—ferumoxytol, heparin and protamine—to form self-assembling nanocomplexes of about 150 nm in size that effectively label cells for MRI. The approach was shown to effectively label three types of stem cells and two types of immune cells.

Targeting mesenchymal stem cells (MSCs), progenitors of osteoblasts, to bone has been a long-standing goal but has had limited success so far. Here, Min Guan and her colleagues deliver a peptidomimetic integrin ligand against integrin α4β1 conjugated to the bone-seeking agent bisphosphonate alendronate as a means of attracting infused and/or endogenous MSCs to the bone surface to stimulate bone formation. The approach was tested in both xenotransplantation and immunocompetent mice, as well as in mouse models of trabecular bone loss induced by aging and estrogen deficiency (ovariectomy).

Zhang and colleagues have developed a new targeted delivery system for RNA interference–based bone anabolic therapy. Using dioleoyl trimethylammonium propane (DOTAP)-based cationic liposomes attached to six repetitive sequences of aspartate, serine, serine (AspSerSer)₆, the system provided selective enrichment of the encapsulated osteogenic siRNA in osteogenic lineage cells at the bone formation surface and the subsequent depletion of the target gene, encoding the bone formation inhibitor casein kinase-2 interacting protein-1 (PLEKHO1, also known as CKIP-1), leading to the promotion of bone formation in healthy and osteoporotic rats.